KICK THE BOARDS USMLE STEP 1BIOCHEMISTRY Prepared by Dr. Irfan Mir KICK THE BOARDS. USMLE STEP 1 BIOCHEMISTRYPrepared by Dr. IRFAN MIR ONLY FOR EDUCATIONAL PURPOSE.1 NOT FOR COMMERCIAL USE OR SALE BIOCHEMISTRY ACID BASE BALANCE :-K1 * HA (acid a proton donor) H+ (proton) A(conjugate base)eg. H2CO3H++ HCO3 K-1 * Remember K1 is forward rate where as K-1 is reverse rate . * pKa :- is the point where forward rate and reverse rate is equal or when acid and base concentration is exactlyequal or a point where proton has been removed from the titrable group. * PI (Isoelectric Point) is the point at pH where sum of the +ve charge is equal to the sum of the -ve charge wherenet charge is 0. * Pka express the strength of acid where as pH measures the acidity of solu. * A Strong acid has a pKa of 2 or less where as Weak acid has a pKa of 10 or more. * A Neutral solu has pH of 7 and normal pH of Blood is 7.4 ( 7.37 - 7.42 ). * Max Buffering capacity occur at pH equal to Pka. * Buffering region extends 1 pH unit above or below the Pka. * Blood at normal pH (7.37 - 7.42) contains * HCO3 --23 - 25 mM * CO2 -- 38 - 48 mmHg * Metabolic acidosis (pH 7.22) showsHCO3--12 mmHg. * Metabolic alkalosis (pH 7.65) showsHCO3--48 mM * CO2-- 30 mmHg*CO2-- 45 mmHg * Respiratory acidosis (pH 7.26) showsHCO3 --26.5 mmHg* Respiratory alkalosis (pH 7.7) showsHCO3-- 22 mM * CO2 --60 mmHg* CO2-- 20 mmHg * In metabolic acidosis HCO3 + as a consequence of the addition of strong acid to ECF. * In respiratory acidosis CO2 | as a consequences of hypoventilation. * In metabolic alkalosis the HCO3 | as a consequence of excessive acid loss ( eg. vomit ) or addition of base ( egantacids ). * In respiratory alkalosis the CO2 + as a consequences of hyperventilation. * In acidosis pH fall 7.45. * In DM type I Oxidation of fatty acid | excessive production of Acetoacetic Acid and 3 Hydroxybutyric Acidand Acetone which are known as Ketone Bodies. * Acetoacetic acid and 3 Hydroxybutyric acid dissociate at body pH and release H+ leading to metabolicacidosis. (on other hand heart and brain use Acetoacetate as alternate fuel). * The combination of high blood level of ketone bodies & metabolic acidosis is called Ketoacidosis which resultinto dehydration, lethargy, vomiting followed by drowsiness and coma.TX is insulin & in severe cases IV NaHCO3. --------------------------------------------------------------------------------------- * o Carbon of each AA is attached to 4 different groups & therefore is CHIRAL or OPTICALLY ACTIVE CARBON ATOM but glycine is exception because it contain two H groups (so is optically inactive and have no side chain). * All AA contain Amino group except Proline in which Amino Group is not present instead there is Imino Group. * When COOH+ (carboxyl group) lose proton it convert into COO(carboxylate group) at physiologic pH. * At physiologic pH 7.4 all AA have both a -ve charge ( COO) Carboxylate Group and +ve charge ( NH3+)Amino Group so called Dipolar ion or Zwitterion.KICK THE BOARDS. USMLE STEP 1 BIOCHEMISTRYPrepared by Dr. IRFAN MIR ONLY FOR EDUCATIONAL PURPOSE.2 NOT FOR COMMERCIAL USE OR SALE * ALIPHATIC -R- Group are non polar, uncharged & hydrophobic group it includes Valine, Alanine, Leucine,Isoleucine, Proline. * HYDOXYL containing -R- Group are mildly polar, uncharged & hydrophilic it includes Serine and Threonine. * SULFUR containing -R- Group include Cysteine and Methionine. * AROMATIC -R- Group are non polar include Phenylalanine, Tyrosine, Tryptophan, * CARBOXL containing -R- Group are carboxylate it include Aspartic acid and Glutamic acid and AmideAsparagine, Glutamine. * BASIC -R- Group include Lysine, Arginine, & Histidine. They all are proton acceptor. * Conformation of protein is determined by rigid planar nature of the Peptide Bond also nature and arrangement of R group. * Stereoisomer (Optical isomers or Enantiomers) define mirror image of same AA ( L form and D form ). * Most drug are either Weak Acid or Weak Base. Drug passes through mem more rapidly if it is uncharged. * HANDERSON HASSEL BATCH Eq can be used to calculate pH of solu containing Weak Acid after addition ofStrong Acid or Base. It can also demonstrate buffering point of a solu following the addition of Acid and Base. pH =pka + log [ A] Ka =dissociation constant of acid [ HA ]HA =weak acid A=conjugate base. * Chaperone are Polypeptide chain binding protein act as catalyst by increasing the rate of final stage in folding protein. * Secondary Structure of protein are :- * o Helix are polypeptide Helices it contain 3.6 AA per turn. Extensive Hydrogen Bonding Stabilizes its structure.(proline, glycine & asparagine are seldom found in o Helixthey are helix breaker) * | Sheet (| Pleated Sheet) form 2 or more peptide chain, arranged in parallel & antiparallel fashion. It alsoinvolve Hydroge Bonding. * | Band (Reverse turn) reverse the direction of polypeptide chain it contain proline and glycine. Structurestabilizes by Hydrogen and Ionic Bonding. * Non Repetitive Secondary Structure :- protein organized in repetitive structures as o helix and | sheet. * Super secondary structure protein are constructed by all. (eg. o helix, | sheet & non repetitive 2 structure) * Tertiary Structure Protein :- are three dimensional protein. Disulfide Bond b/w Cysteine residues may stabilize3 protein. Also Hydrophobic interaction, hydrogen bonding, ionic interaction stabilizes the 3 structure protein. * Quaternary structure protein :- hasmore than one peptide chain in protein subunit. ----------------------------------------------------------------------------------------- * Globular protein are polar and soluble in aqueous solu with hydrophobic side chain (R group) inside andhydrophilic R group facing out side in soluWhere as mem protein have theirhydrophobic R group facing on the surface (out side). * In Sickle cell anemia ( Homozygous genotype SS ) valine replaces the glutamate at the position of 6 of | chainof normal Hg A (hgb A)to form mutant sickle cell hemoglobin (hgb S). * In Sickle cell trait (heterozygous genotype AS) one normal (hg A) & one abnormal hemoglobin (hgS) is present intheir RBC. The condition is usually asymptomatic with no anemia. Episodes of hematuria is due to sickling of RBCsin adrenal medulla and is self limiting. * Ascorbic acid is required for hydroxylation of proline and lysine during post transitional process of collagentogether with enz hydroxylase, O2, Fe2+.(Hydroxyproline form interchain Hydrogen bond that stabilize collagen triple helix). Ascorbic acid +missing Hydrogen bond and weaken collagen Scurvy and impaired wound healing. * Ascorbic acid is also required to maintain iron in its active oxidative Fe2+state. KICK THE BOARDS. USMLE STEP 1 BIOCHEMISTRYPrepared by Dr. IRFAN MIR ONLY FOR EDUCATIONAL PURPOSE.3 NOT FOR COMMERCIAL USE OR SALE * A G (free energy change) is the quantity of energy from reaction that is available to do work. * A H (Enthalpy change) is the amount of heat generated or absorbed in the reaction. * A S (Entropy change) measure the change in the randomness or disorder of the system. * A G is the standard Free Energy Change. * A G is the energy required to scale the energy barrier and form the transition state. * The greater AG the lower the rate of reaction where as lower AG faster the rate of reaction. * MICHAELIS - MENTEN EQUATION predict how velocity is related to substrate conc if enz conc is held constant. V = Vm [S] /Km + [S].* S is the substrate, Km is the Michaelis constant, Vm is velocity max. Km is (roughly) an inverse measure of the affinity or strength of binding between the enzyme and its substrate. The lower the Km, the greater the affinity . * LINEWEAVER - BURK EQUATION is the form of Michaelis - menten Eq which is some time known as Double Reciprocal Eq. 1= Km + [ S ] =Kmx 1 + 1 . V Vm [ S ]Vm SVm * COMPETETIVE INHIBITORS are substrate analogs that compete with substrate for the active site of enz and resultinto low enzymatic activity ( Km is higher , Vm remain same ). + On the Line Weaver - Burk Plot the slope is increased, the X is intercept has similar absolute value & Y intercept is Unchanged . * NON COMPETETIVE INHIBITORS bind at the site different from the active site ( Vm is lower , Km is unchanged ) + On Line Weaver - Burk Plot the slope is increased, X intercept is unchanged and Y intercept is larger. * UN COMPETETIVE INHIBITOR bind only to the enz - substrate complex (ES complex) both Vm and Km is different. + On Line Weaver - Burk Plot the line are parallel.

* Allosteric regulation :- A low molecular wt effectors binds to the enz at the specific site other than the active siteand alter its activity. Allosteric enz have more than one sub unit & more than one active site. Eg HexokinaseCatalyzes the first reaction in glycolysis (Hexokinase convert glucose glucose 6 ph). Glucose 6 ph allostericallyinhibit hexokinase to ensure glucose 6 ph supply is in balance (thus prevent over accumulation of glucose 6 ph). * Enz can also be regulate by repression of enz synthesis by altering gene expression, phosphorylation etc. * Methanol is oxidized to formaldehyde and formic acid by enz alcohol dehydrogenase. (formaldehyde causeblindness). * Ethylene Glycol is oxidized to glyceraldehydes, oxalate, and lactate by enz alcohol dehydrogenase. (oxalatecrystal deposition in kidney cause kidney failure). * Ethanol is the competitive substrate for enz alcohol dehydrogenase and it displaces the methanol and ethyleneglycol from active site of enz (alcohol dehydrgenase) . thats why ethanol is the Tx of methanol and ethyleneglycol poisoning.--------------------------------------------------------------------------------------------------- KICK THE BOARDS. USMLE STEP 1 BIOCHEMISTRYPrepared by Dr. IRFAN MIR ONLY FOR EDUCATIONAL PURPOSE.4 NOT FOR COMMERCIAL USE OR SALE CARBOHYDRATE METABOLISM & OXIDATIVE PHOSPHORYLATION * Phosphoanhydride Bond are very high energy bond where as Phosphate ester Bond are very low energy bond. * Kreb Cycle (Tricarboxylic acid Cycle or Citric Acid Cycle) located in mitochondria & is the final common pathway of Oxidative metabolism. * Glucose Catabolism produce Pyruvate which yield Acetyl CoA which condensed with OAA (oxaloacetate) tobegin the Citric Acid Cycle. Fatty Acid also generate Acetyl CoA via | Oxidation to join Citric Acid Cycle. (someAA also degraded to acetyl CoA) * Succinyl CoA, ATP and NADH are citric Acid cycle inhibitor (regulator). * TCA cycle provide 11 ATP molecule per cycle by producing 3 NADH &1FADH.TCA cycle also produce one equivalent of high energy phosphate as a GTP. (Always remember 1 NADH generates 3 ATP and 1 FADH generates 2 ATP).* Remember ATP allosterically inhibit acetyl CoA. where as NADH & ATP inhibit Isocitrate allosterically and SuccinylCoA & NADHinhibit o ketoglutarate allosterically. They all are TCA cycle regulator. * NADH release by conversion of Isocitrate to o ketoglutarate than o ketoglutarate to succinyl CoA & form Malateto Oxaloacetate by respective dehydogenase. * GTP is generated while succinyl CoA convert into succinate by enz succinyl CoA synthatase. later succinate tofumerate reaction by enz succinate dehydrogenase produce FADH2.

Electron transport & Oxidative phosphorylation: * Mostly ATP generate from mitochondrial oxidative phosphorylation in Aerobic cell. When ATP get utilized itproduce ADP and Pi(Piis inorganic phosphate). subs that inhibit Electron transport chain inhibit ATP syn. * In Electron transport chain system electron pass from NADH or FADH2 to ultimately reduce O2 and produce H2O.This produces ATP by oxidation phosphorylation which involve 5 complexes. 1. Complex I uses NADH dehydrogenase to convert NADH to NAD.(Inhibited by amobarbital, Hotenone (insecticide), Picricidin (antibiotic)). 2. Complex II uses succinate dehydrogenase to convert FADH2 to FAD. (It is inhibit by Antimycin (antibiotic)). 3. Complex IV uses cytochrome oxidase convert O2 to H2O. (it is inhibited by Cyanide, Hydrogensulfide, CO). * ATP synthatase complex (complex V) :- H+ion pass back into the matrix through complex V and in doing sodrive sys of ATP. Subs like uncoupling protein (Thermogenin, certain chemicals) caries H+across the innermitochondrial mem with out going through complex V (with out synthesizing ATP) are normally dissipated intoheat and maintain body temp. * Subs like uncoupling protein (Thermogenin), chemicals which carry H+ across the inner mitochondrial mem without going through complex V ( with out synthesizing ATP ). This energy dissipated into heat to maintain bodytemp. Carbohydrate Metabolism:* Monosacchrides (Glucose, Fructose) are absorbed through carrier mediated transport and stored as glycogenor converted to triglycerides or produce energy via oxidation. * Uridine diphosphate - Glucose (UDP-G) is activated substrate for gluconeogenesis. The enz glycogen synthatase add glycosyl unit to non reducing end in o - 1,4 linkage & amylo transgycolase create branches with o - 1,6linkage. KICK THE BOARDS. USMLE STEP 1 BIOCHEMISTRYPrepared by Dr. IRFAN MIR ONLY FOR EDUCATIONAL PURPOSE.5 NOT FOR COMMERCIAL USE OR SALE * In Glycogenolysis enz phosphorylase release on unit glucose - 1- ph at a time from non reducing end. * Glucose - 1 - ph convert into Glucose - 6 - ph by phosphoglucomutase. * Glucagon on liver andEN on liver and muscle stimulate glycogenolysis and inhibit Glycogenesis vs.Insulin stimulate glycogenesis and inhibit glycogenolysis via dephosphorylation in muscle liver & adipose tissue. * Retina has high rate of oxidative metabolism any disturbance Retinal degeneration and blindness. * In Anaerobic glycolysis glucose convert into 2 lactate and 2 ATP. vs. Where as in Aerobic glycolysis glucoseproduce 6CO2, 6H2O, 36 - 38 ATP.( Glucose +6O2 6CO2 +6H2O +36 - 38 ATP ) * Anaerobic glycolysis is characteristic of muscle after prolong exercise and Hypoxia. Where enz lactatedehydrogenase convert pyruvate to lactate. * Phosphorylation is the 1st step in Glycolysis in which glucose convert to G -6 -ph by enz hexokinase & glucokinase Hexokinase present in most tissues where as glucokinase present in liver and pancrease. * Hexokinase inhibit by (regulate) G - 6 - phwhere asGlucokinase inhibit by (regulate) F - 6 - ph to preventaccumulation of G - 6 - ph and F - 6 - ph. ----------------------------------------------------------------------------------------------- * Gluconeogenesis synthesize glucose from non carbohydrate precursor, (for eg. lactate & alanine) primarily inliver, kidney. * Gluconeogenic substrate are lactate, Pyruvate, Glycerol, and subs that can be converted to OAA(oxaloacetate) than reverse glycolysis occur to produce glucose. * Cori Cycle describe the shuttling of Gluconeogenic substrate b/w muscle and liver ( which is lactate ). Lactatetransport from muscle to liver where it use in glucose synthesis. than glucose is release into the blood. * | F, 2,6 Bph ( fructose 2,6 biphosphate ) and AMP and + Glucagon inhibit Gluconeogenesis and favor Glycolysis where as+ F, 2,6 Bph and | Glucagon promote Gluconeogenesis. * In starvation glucagons is high and in fed state glucagons is suppressed. ------------------------------------------------------------------------------------------------ * Pentose Phosphate Pathway Function as alternative form of glycolysisor may be route of complete oxidation of glucose. * Pentose ph Pathway ( HMP Shunt ) : + Non Reversible Oxidative Reaction ----------------- Generate NADPH + Reversible Non Oxidative Reaction portion rearrange sugar and reenter in glycolysis pathway . * Ribose - 5 - ph which is needed for nucleotide syn can be form from G - 6 - ph by either arm of pentosephosphate Pathway. -------------------------------------------------------------------------------------------------- * Enz Sucrase convert Sucrose (sugar from cane sugar)into glucose and fructose. * Enz lactase convert Lactose (milk sugar) into glucose and glactose. * Fructokinase deficiency (Failure of F-1-ph conversion from Fructose) essential fructosuria (benign disorder). * Fructose -1- ph aldolase deficiency (Failure of Dihydroxyacetone ph & glyceraldehydesconversion from F-1-ph) severe hypoglycemia after fructose or sucrose ingestion(fructose intolerance). * Lactase deficiency (some time in adult life) results into milk intolerance, bloating, Flatulence, and diarrhea. * Galactose kinase deficiency ( Failure of Glactose -1- ph conversion from gatactose ) mild galactosemia andearly cataract. * Galactose -1- ph uridyl transferase deficiency ( Failure of Glucose -1- ph conversion from Galactose -1- ph ) severe galactosemia which result in growth failure, mental retardation, and even early death. KICK THE BOARDS. USMLE STEP 1 BIOCHEMISTRYPrepared by Dr. IRFAN MIR ONLY FOR EDUCATIONAL PURPOSE.6 NOT FOR COMMERCIAL USE OR SALE GLYCOGEN STORAGE DIS : 1. Von Gierkes dis (type I) caused by glucose - 6 - ph deficiency glycogen accumulation in liver & kidney with resultant Failure to thrive, hypoglycemia, ketosis, lipidemia and hyperurecemia. 2. Pompes Dis (type II) caused by o 1,4 glycosidase deficiency glycogen accumulation in lysosomes in everyorgan tissuewith resultant Cardiac and respiratory failure and death before age 2. 3. Coris Dis (Type III) caused by debranching enz deficiency short outer branched glycogen accumulation inliver & muscle which cause Similar symptoms like type I but milder. 4. Anderson Dis (Type IV) caused by branching enz deficiency long outer branched glycogen accumulation inliver & spleen with resultant Liver chirrosis and death before age 2. 5. McArdle Dis (Type V) caused by Phosphorylase deficiency glycogen accumulation in the muscle painfulmuscle cramps with exercize. ---------------------------------------------------------------------------------------- LIPIDS * Lipids :- 1. Fat: (Triacylglycerol, Triglycerides) are major fuel store of body. (excessive dietary fat is stored as Triacylglycerol) 2. Phospholipids: are cell mem lipid and part of transport lipid in blood. 3. Sphingolipid: are component of membranes. 4. Cholestrol: are component of cell mem, precursor of bile acid, bile salt and steroid hormones. Triglyceride Composition :- Stearic acid, Oleic acid, Palmitic acid, Glycerol (synthesize by o glycerophosphate) * Chylomicron consist of :- + 87% Triglyceride -- TG hydrolysed to FFA and Glycerol by lipoprotein lipase on the surface of capillaryEndothelium ofmuscle and adipose tissue. + 9% Phospholipids + 3% Cholesterol + 1% Apoprotein -- make chylomicron suspension able in lymph fluid there by prevent attachment to lymph wall. * This is Apoprotein that make chylomicron suspension able in lymph fluid and prevent attachment to lymph wall. * Lipoprotein lipase present in adipose tissue and liver, when blood passes through them the most Chylomicron present in the blood is removed by endothelial cells which are rich in lipoprotein lipase. * Lipoprotein lipase hydrolyze the Triglycerides (TGR) into Fatty acid (FA) & Glycerol than FA enter into fat cell orliver resynthesize TGR to store into the cell. Lipoprotein lipase also hydrolyze the phospholipids. * Hydrolysis of Triglycerides is stimulate by 1. + o glycerophosphate2. Hormones like ACTH, glucocorticoids, EN, norEN, TH. * Types of Lipoprotein :- 1. VLDL(| Triglyceride, moderate Cholesterol and Phospholipids) 2. IDL (| cholesterol and phospholipids , + Triglyceride) 3. LDL(| cholesterol and phospholipids) 4. HDL (| protein, + cholesterol and phospholipid)

All function as transporter of Triglycerides, cholesterol or phospholipids & all lipoprotein synthesize in liver except small intestine also synthesize small HDL. * Triglycerides are of two types ----- 1. fatty acid, 2. glycerol (which enter the cell carrier is carnitine than into mitochondria to produce FA Acetyl CoA for | oxidation process) * One Acetyl CoA when split from fatty acid release 4 H atom used in ATP formation than Acetyl CoA enter into Citric acid cycle here again it degraded into CO2 and H atom. H than oxidized by chemosmotic Oxidation system of mitochondria. * Two Acetyl CoAcondensed to form Acetoacetic acid in liver which is than transported to peripheral tissue fromliver because only small fraction of fatty acid is used by liver to fulfill its metabolic need. KICK THE BOARDS. USMLE STEP 1 BIOCHEMISTRYPrepared by Dr. IRFAN MIR ONLY FOR EDUCATIONAL PURPOSE.7 NOT FOR COMMERCIAL USE OR SALE Acetoacetic acid (in liver)| Hydroxybutyric acid acetone (in liver) cell (peripheral)reverse action tomake Acetyl CoAused in citric acid cycle. * Fatty acid must | oxidized to form Fatty Acetyl CoA which enter in the Citric acid cycle where it oxidized to CO2and H2O. Each cycle of | oxidation generate 5 ATP via electron transport system and 11 ATP, 1 GTP & OAA viaCitric acid cycle. * Ketones Body are1. Acetoacetic acid (keto acid)2. | Hydroxybutyric acid3. Acetoac (Acetone)* Ketosis occur when ketone bodies conc in plasma rises usually due to starvation or in DM or in person with entirefat diet. * Synthesis of Triglycerides from Carbohydrates:- synthesis occur in liver and adipose tissue. Carbohydrate acetyl CoA malonyl CoA and NADPHfatty acidTriglycerides Acetyl CoA carboxylase ( carboxylation) glycerol * Regulation of energy release from Triglycerides:- | o glycerophosphate (product of glucose metabolism)inhibits fatty acid release for energy and | fat stores. + o glycerophosphate stimulate fatty acid release for energy. Hormones-- + insulin, TSH, CTH, ACTH, EN, Nor EN. * Phospholipids are1. Lecithins 2. Cephalins3. Sphingomylins (All body cells synthesis phospholipids but liverhas more ability (90%)). * Phospholipids always contain one or more fatty acid and one phosphuric acid radical and usually containnitrogenous base. * Synthesis of phospholipids | when Triglycerides |. * Chemicals also contain phospholipids like choline (from nitrogenous base) and inositol (from cephalin).Phospholipids also present inlipoprotein, thromboplastin, sphingomyelin in CNS, structural element. * Cholesterol are highly fat soluble & slightly water soluble. In Plasma 70% of cholesterol are in the form of esters. * Endogenous cholesterol synthesize in liver by 3 hydroxyl 3 methyl glutaryl CoA reductase. (| blood cholesterollevel inhibit that enz). * | saturated fat diet | acetyl CoA in liver | cholesterol production. * | unsaturated fatmoderate production of cholesterol. Function of Cholesterol :- Cholic acid syn bile salt. Syn of hormones ACTH, Progesterone, testosterone, prevent water evaporation from skin.

* Mouth lipase hydrolysed TG, than DAG & FFA (free fatty acid) is produced in stomach later Bile salt emulsifiedlipids in duodenum. * In small intestine emulsified fat hydrolyzed by pancreatic lipase, Phospholipase A & Cholestrol estrase. * Mixed Micells formation contain fatty acid, DAG, monoacylglycerol, phospholipids, cholesterol, Vitamin ADEKand bile acid. * Medium chain Fatty acids passed into portal vein vs.long chain Fatty acid reincorporated into TG, than TG incorporated into chylomicron which pass into lymphatic. * Lipids are transported in Plasma via lipoprotein. * Exogenous lipid transport in plasma via chylomicron vs. endogenous lipid release as VLDL. * FFAs release by adipose tissue absorbs by muscle for oxidation. * In adipose tissue lipase is activated by glucagons and EN to hydrolyzed TGs into FFAs. * VLDL (endogenous TGs) are hydrolyzed to FFAs and Glycerol by lipoprotein lipase and yield LDL. * LDL cholestrol inhibits HMG CoA Reductase, that is the rate limiting enz in cellular cholestrol syn. LDL also down regulate LDL receptor synthesis. * HDL synthesized by liver serve as a reverse cholestrol transport. KICK THE BOARDS. USMLE STEP 1 BIOCHEMISTRYPrepared by Dr. IRFAN MIR ONLY FOR EDUCATIONAL PURPOSE.8 NOT FOR COMMERCIAL USE OR SALE * FA must activated as their acetyl CoA thioesters & than Acetyl CoA & oxidized to CO2 & H2O by | Oxidation togenerate ATP. * Acetyl CoA & Malonyl CoA synthesize fatty acid synthase complex in cytosol Fatty acid syn. (Fatty acid synincreases when cell is starving). AcetylCoA is transported from mitochondria to cytosol by the citrate- malate-pyruvate shuttle for Fatty acid syn. * Fatty acid yield propionyl CoA as final product of | Oxidation than propionyl CoA carboxylated to Succinyl CoA(in three reactions) to enter the Citric acid cycle.(remember Fatty acid enter the TCA cycle as a Acetyl CoA and Succinyl CoA) * Ketogenesis occur when some of the Acetyl CoA is metabolized to Acetoacetate and | Hydroxybutarate in liver. Glucose starved brain can use acetoacetate for fuel. Heart can also utilize acetoacetate for energy (at theexpense of succinyl CoA). * Linoleic acid serve as a precursor for arachidonic acid. (remember arachidonic acid is required for the syn of PG, Thromboxane & eicosanoids) Remember Linoleic acid and Lenolenic acid are essential fatty acid diet must contain it. * Steatorrhea (due to mal absorption of fat) caused by Celiac dis, Abetalipoproteinemia, Non Tropical Sprue, Inflammatory Bowel dis. * Hyperlipidemia can be TX with HMG CoA Reductase inhibitor ( eg. all statins ) which reduces endogenouscholesterol syn. HMG CoA Synthatase HMG CoA Reductase (Multiple step process) Acetyl CoAHMG CoA Mevalonic acid cholesterol Acetoacetyl CoAfeed back inhibition

* In mixed hyperlipidemia there is a danger of Acute Pancreatitis. It is characterize by over production of VLDL,Defective lipolysis of VLDL triglycerides (Endogenous) and chylomicron (Exogenous). (both serum TGR andcholesterol is +) * The syn of Sphingolipids begins with with palmityl CoA and serine. * The no of sphingolipid storage dis may occur due to deficiency of one of the lysosomal enz. (sphingolipidnormally digest in lysosomes) ** Lysosomal (sphingolipids) Storage dis :-(Lysosomal enz deficiency) 1 Tay-sachs Dis : ARD, Hexoaminidase Deficiency Ganglioside GM2 accumulationmental retardation,blindness, cherry red spot on macula & death by age 3. 2 Gauchers Dis : ARD, Glucocerebrosidase deficiencyglucocerebroside accumulationhepatosplenomegaly, bone erosion, mental retardation. 3 Nieman- Pick Dis : ARD: Sphingomyelinase deficiency Sphingomyelin accumulation hepatosplenomegaly,mental Retard. 4 Fabrys Dis : X - linked dis, o galactosidase A deficiencyCeramide trihexoside accumulation Angiokeratoma like skin lesion, kidney failure, lower extremities pain. 5 Krabbes Dis (Globoid Cell Leukodystrophy) : Galactosylceramide | galactosidase defGalactocerebrosideaccumulationmental retardation,absent myelin. ----------------------------------------------------------------------------------- * Deamination is first step in metabolizing surplus AA it yield o Ketoacid & NH4+ (ammonium ion) byAminotransferase (Transferases) and glutamate dehydrogenase. KICK THE BOARDS. USMLE STEP 1 BIOCHEMISTRYPrepared by Dr. IRFAN MIR ONLY FOR EDUCATIONAL PURPOSE.9 NOT FOR COMMERCIAL USE OR SALE * In most tissue NH4+ and glutamate becomes glutamine which than enter the and convert back to NH4+andglutamate. * Similarly in skeletal muscle Glutamate dehydrogenase and glutamate - Pyruvate aminotransferase can lead toincorporation of NH4+ into Alanine. Then in liver transamination results alanine back into Pyruvate and NH4+. * NH4+(Ammonium ion) is toxic to human body specially CNS, thats why body convert NH4+to urea in liver viaurea cycle . Than urea excreted in urine NH4+ + GlutamateGlutamine( in most tissue ) NH4++Glutamate( in liver ) Glutamine synthatase Glutaminase * UREA CYCLE :- * When any of enz in urea cycle +( sp Carbomoyl phosphate synthase I& Ornithinine transcarbomoylase ) Hyperammonemiamental retardation, seizure , coma and death.TX involve Restriction of Dietary protein with intake of ketoacid& essential AA and also Benzoate & phenylacetate to provide alternative pathway for ammonium excretion. * There are 9 Essential AA.PHILL MTVT ( Leucine, Isoleucine, lysine, Valine, phenylalanine, Methionine, Threonine,histidine, Tryptophan.) -------------------------------------------------------------------------------------------------------- * Acetyl CoA family also called Ketogenic AA Family are Leucine, Isoleucine, Lycine are ketogenic AAwhereas Phenylalanine, Tryptophan, Tyrosine are Ketogenic and glucogenicAA. * Tryptophan use in serotonin syn & Histidine use in histamine syn. * Methionine use in cysteine syn & methylation reaction. * Phenylalanine metabolize to tyrosine by enz phenylalanine hydroxylase later tyrosine is used in syn of EN, Nor EN,Triiodothyronine, Thyroxine, Dopamine, melanin. * Phenylketonuria (PKU) : phenylalanine Hydroxylase or dihydropteridine Reductase Deficiency Hyperphenylalaninemia mental retardation, CNS damage, Hypopigmentation.(alternative pathway of phenylalanine breakdown phenylketones in blood & urine). TX isphenylalanine free diet but dietary tyrosine becomes essential AA. * Albinism --- Tyrosinase deficiencyabsent melanin, photophobia, susceptibility to sun burn & skin carcinoma. * Homocystinuria occur due to 1. Cystathionine synthatase deficiency --------2. Reduced affinity of Cystathionine synthatase for its coenz Pyridoxal phosphate (PLP) ----- respond to mega doses of pyridoxine. 3. N-Methyltetrahydrofolate homocysteine methyltransferase Deficiency. 4. Vitamin B12 deficiency -----------respond to B12 supplement * Homocysteinuria results in dislocation of optic lens, mental retardation, Osteoprosis, Atherosclerosis, thromboembolism.TX for those who do not respond to Vit therapy is diet low in methionine and BETAINE (NNN trimethylglycine) supplement as a alternative methyl group donor which lack sulfur group.(because sulfur frommethionine can be used in cysteine syn) . * Maple Syrup Urine Dis ----- branched chain 2 ketoacid decarboxylase defsevere brain damage and death in first yr of life. Branched chain ketoacid derived from leucine, isoleucine & valine appear in urine giving it maplesyrup like odor.TX -- few cases respond to megadosis of thiamine B1 other wise diet low in branched chain AA are given. * Histidinemia ----- deficiency of histidine o Deaminasemental retardation and speech defect may occur butis rare. (| histidine and its metabolites in urine) .TX is usually not indicated. ------------------------------------------------------------------------------------ KICK THE BOARDS. USMLE STEP 1 BIOCHEMISTRYPrepared by Dr. IRFAN MIR ONLY FOR EDUCATIONAL PURPOSE.10 NOT FOR COMMERCIAL USE OR SALE * A nucleotide is a phosphorylated nucleoside. (or nucleoside with one or more phosphate group is nucleotide). * Nucleotide contains 1. Base ----------------------------------------- 1. Purine --------------------------------------------- + Adenine ( In DNA & RNA ) 2. Pyrimidines --- +Cytosine ( In DNA & RNA ) + Guanine ( In DNA & RNA ) +Thymine ( In DNA only ) 2. Sugar -------------------------------v Ribose (RNA) +Uridine ( In RNA only ) v Deoxyribose (DNA) 3. Phosphate Group * Nucleotide is 1. Substrate for DNA or RNA . 2. Carrier for high energy group ( ATP, UDP, acetyl CoA, Methyl group etc ). 3. Component of Coenz (NAD, NADP, FAD, CoA) 4. Regulating molecules ( cAMP, cGMP) * Deoxyribose Sugar found in DNA which has H+ on 2nd position vs. Ribose sugar found in RNA which has OH at 2nd position. * Thyamine Base found in DNA only & has CH3 in Base ring vs. Uracil Base found in RNA only & has no CH3 inBase Ring. * Purine nucleotide Syn begins with syn of 5' phosphoribosyl -1 - pyrophosphate (PRPP) by PRPP synthatase. (IMP,AMP & GMP inhibit PRPP synthatase)vs. Syn of Pyrimidine starts with syn of carbomoyl phosphate by carbomoylphosphate synthatase II. * Salvage Purine Bases is another way to synthesize purine nucleotide it involve 2 enzs HGPRTase (inhibit by IMP &GMP) and Adenine Phosphoribosyl transferase (inhibit by AMP). vs.Salvage of Pyrimidine is accomplishedby enz pyrimidine Phosphoribosyl transferase. * One of the product of purine degradation is uric acidvs.pyrimidine degradation produce | AA, CO2, NH4+. * In purine degradation Hypoxanthine and xanthine are oxidized to uric acid by enz xanthine oxidase. * In pyrimidine degradation the product | Aminoisobutyrate (which originate exclusively from thyroninedegradation) is clinically imp. because urinary | Aminoisobutyrate is an indicator of DNA turnover followingchemotherapy, radiation, cancer. * Hereditary Orotic Aciduria -------- deficiency of Orotate phosphoribosyl transferase & OMP decarboxylase(nucleotide metabolism enz) retarded growth and severe anemia. TX is feeding synthetic cytidine or uridinewhich supplies the pyrimidine for RNA and DNA synthesis. (UTP from these synthetic nucleoside act as feed backinhibitor ofcarbomoyl phosphate Synthatase II so shut down orotic acid syn)* Purine Nucleoside Phosphorylase deficiency | level of purine nucleoside with + uric acid formation & impairedT - cellfunction. * Severe Combined Immune Deficiency (SCID) -- adenosine deaminase deficiency T & B cell dysfunction. TX:gene therapy. * Lesch Nyhan $ --- HGPRTase deficiency (or absence of salvage enz) which results in no salvage of Hypoxanthineand Guanine which further + the IC IMP and GMP for that reason de novo Pathway is not properly regulated onother hand IC PRPP |which stimulate the de novo pathwayexcessive purine syn, hyperuricemia ( increase uric acid formation ), orange color stool, severeneurologic problem, including spasticity, mental retardation, selfmutilation. TX by Allopurinol which + sodium urate crystals but it does not ameliorate the neurologic symptom * Hydroxyurea (anti cancer agent) inhibit nucleoside diphosphate reductase which further inhibit ribonucleotideconversion to deoxyribonucleotide. * Aminopterin and methotrexate inhibit dihydrofolate reductase so inhibit conversion of dihydrofolate totetrahydrofolate thus inhibit thymidylate syn. * Anti cancer agent Fluorodeoxyuridylate inhibit thymidylate synthatase, the enz that convert dUMP to dTMP. KICK THE BOARDS. USMLE STEP 1 BIOCHEMISTRYPrepared by Dr. IRFAN MIR ONLY FOR EDUCATIONAL PURPOSE.11 NOT FOR COMMERCIAL USE OR SALE * Gout result from disorder of purine metabolism (de novo syn) may be due to mutation in PRPP synthatase with lossof feed back inhibition by purine nucleotides with resultant over production of purine and uricacid. (a partialHGPRTase deficiency may develop which lead to less consumption of PRPP with resultant more purine syn) * Uric acid is not soluble in body fluid it deposit in joint as sodium urate crystals also in muscle & kidney whichcause arthritis & renal damage. TX of gout is Allopurinol (Xanthine oxidase catalyse oxidation of allopurinol toalloxanthine which is potent inhibitor of xanthine oxidase ).Allopurinol | level of hypoxanthine & xanthine which is more soluble in body fluid than uric acid & do not formcrystals. * Sexpndary reasons of gout include Myeloproliferative $, Chronic renal failure, Lesch Nyhan $, radiation & cancerchemotherapy. --------------------------------------------------------------------------------- * Daily energy need ( Kcal/lb ) is highest in infancy ( 55Kcal/lb ) & lower as age Inc 13 - 15 Kcal/lb in adult (per day). * Carb--- provide 4 Kcal/gm. ( carbohydrate should comprise 50 - 60 % of calories intake ) Protein--- provide 4 Kcal/gm ( protein should comprise 10 - 20 % of caloric intake ) Fat--- provide 9 Kcal/gm ( fat should comprise >30 % of caloric intake ) Alcohol--- provide 7 Kcal/gm. * Inadequate carbohydrate intake ketosis by excessive break down of tissue protein (wasting), loss of cation(Na+), & dehydration. * Essential fatty acid (EFA) are linoleic acid & linolenic acid. (remember linoleic acid is the component ofArachidonic acid) * Nitrogen balance is the difference b/w nitrogen intake (primarily as protein) and nitrogen excretion undigestedprotein in feces, and urea & ammonium ion in urine. In healthy person Nitrogen intake is equal to excretion. * Nitrogen balance :- + +ve Nitrogen Balance -- in which intake exceeds excretion, it occur when protein requirement |eg as in pregnancy, lactation, growth, recovery from surgery, trauma & infection. + - ve Nitrogen Balance -- In which excretion exceeds intake. Occur during metabolic stress, +protein intakewhen essential AA is missing with insufficient food intake including both calories & protein. * RDA for protein is 0.8gm/kg or 60gm for 75 kg man. * Marasmus is caused by starvation with insufficient food intake including both calories and protein. * Kwashiorkor is starvation with edema due to diet deficient more in protein than total calories. * In Marasmus ketogenesis provide fuel for brain and cause muscle wasting and depleted subcutaneous fat vs.In Kwashiorkor liver is enlarged due to fatty infiltration and depleted subcutaneous fat severely, edema, flakypaint dermatitis etc* BMI 20 is desirable, 25 is over wt, where as 30 and over is obese. 20 % of adolescent and 30 % of adults are overwt in USA. * Disease associated with obesity are | serum lipid, coronary artery dis. hypertension, Non insulin dependent DM,breast and uterine cancer, gallstone, osteoarthritis, respiratory problem (reduced functional lung vol.). ---------------------------------------------------------------------------- VITAMINS * Thiamine B1is required for proper nerve transmission & a Coenz for pyruvate, o ketogutarate dehydrogenase,transketolase (in pentose ph pathway), branched chain ketoacid dehydrogenase. B1 deficiency lead to beri beri in three stages. 1. Early symptomsare appetite loss, constipation, nausea, peripheral neuropathy, fatigue. 2. Moderate to severe cases Wernicks Korsakoffs $ in alcoholics shows confusion, ataxia, ophthalmoplegia. 3. Severe cases + Dry beri beri : all above symptoms plus wrist and foot drop. + Wet beri beri : symp of dry beriberi + high potency cardiac failure, pul congestion & edema.KICK THE BOARDS. USMLE STEP 1 BIOCHEMISTRYPrepared by Dr. IRFAN MIR ONLY FOR EDUCATIONAL PURPOSE.12 NOT FOR COMMERCIAL USE OR SALE * Riboflavin B2 is converted to oxidation reduction Coenz FAD (flavin adenine dinucleotide) and FMN. Riboflavin Deficiency results into + Angular cheilitis (cracking of the corner of lip) + Glossitis(Red and swollen tongue) + Scaly dermatitis particularly at nasolabial fold and around scrotum. * Niacin B3 (nicotinic acid, niacinamide, nicotinamide) is converted into oxidation reduction Coenz NAD(nicotinamide adenine dinucleotide) and NADP. Niacin can be synthesize from AA tryptophan.Niacin deficiency v mild deficiency results into glossitis v Sever deficiency results into Pallagra (dermatitis, dementia, diarrhea) High doses of niacin (2- 4gm/day) cause vasodialation (rapidly) and lower blood cholesterol and LDL. * Pantothanic acid B5 is component of CoA and phosphopantetheine of fatty acid synthatase. deficiency is rare. * Pyridoxine B6 (pyridoxamine, pyridoxal) pyridoxal ph is Coenz involve in transamination. Pyridoxine def+ mild --- irritability, nervousness and depression + severe --- peripheral neuropathy and convulsion and occasional Sidroblastic anemia. + other symptoms are eczema, seborrheic dermatitis, chapped lips, glossitis, angular stomatitis. High doses of pyridoxine B6 used in homocysteinuria which results from defective Cystathionone -|- synthase. Prolong consumption of B6 lead to toxicity with sensory neuropathy. * Biotin is prosthetic group for carboxylation enz eg pyruvate carboxylase, acetyl CoA carboxylase. Biotin is also synthesize by intestine bacteria. Biotin deficiency dermatitis, hair loss, atrophy of lingual papillae, gray mucous mem, muscle pain, paresthesia, hypercholesterolemia, ECG abnormality. (20 raw eggs ingestion cause biotin deficiency other wise deficiency is rare). * Folic acid serve as a Coenz in one carbon transfer reaction in purine and pyrimidine syn, thymidylate syn,conversion of homocysteine to methionine, serine glycine interconversion. Folic acid : 1. aloblastic anemia (similar to B12 +) due to blocked DNA syn. 2. Neural tube defect as a result of maternal folate deficiency. 3. | Body Homocysteine which cause atherosclerotic heart dis with B6 deficiency. 4. Methotrexate, TMP, Pyrimethamine (antimalerial), Diphenhydrantoin, primidone (anticonvulsant). * Cobalamine B12 is Coenz for methylmalonyl CoA mutase & homocysteine methyltransferase & DNA syn. It foundin meat, sea food and dairy food. B12 Deficiency cause by vegi diet, + gastric intrinsic factor, pancreatic deficiency, crohn's dis, Diphyllobothriumlatum ingestion 1. Megaloblastic anemia (similar to folate) due to block DNA syn, 2. Paresthesia, weakness, some time irreversible CNS & PNS damage. * Ascorbic acid ( vitamin C ) is a coenz for oxidation reduction reaction, including the post translationalhydroxylation ofproline and lysine in maturation of collagen, carnitine synthesis, tyrosine metabolism, andcatecholamine syn. Vitamin C is antioxident and also facilitate iron absorption. Vitamin C deficiency 1. Mild -- capillary fragility, petechiae, + immune function. 2. Severe -- scurvy, + wound healing, osteoprosis, hemorrhage, anemia, teeth may fall out. * Vitamin A (Retinol & Retinoic acid) is an antioxidant it regulatetissue growth & differentiation. 11 - cis - Retinal is a prosthetic group of Rhodopsin (the visual pigment) in Rods and Cones. Body convert | carotine (precursor of Vit A) to retinol (active vit A) to store in liver. Vitamin A def Night Blindness & Xerophthalmia, Progressive keratinization of Cornea, follicular hyperkeratosis (like goose bumps), anemia in the presence of adequate iron store, + resistance to infection, | susceptibility to cancer, impair syn of Retinol binding protein. * Remember Retinal degeneration is cause by Vit E deficiency. KICK THE BOARDS. USMLE STEP 1 BIOCHEMISTRYPrepared by Dr. IRFAN MIR ONLY FOR EDUCATIONAL PURPOSE.13 NOT FOR COMMERCIAL USE OR SALE Vitamin A toxicity Bone pain, scaly dermatitis, hepatosplenomegaly, nausea, diarrhea. (excess | carotine is not toxic). Tretinoin (All Trans - Retinoic acid) & Isotretinoin (13 - cis Retinoic acid) --- TX of Acne. Etretinate ---- TX of Psoriasis. * Vitamin D -- | Blood Ca++ level by stimulating syn of Ca++binding protein in intestinal mucosa. In combination with PTH cause bone demineralization & renal Ca++ reabsorption | blood Ca++. UV radiation convert 7- dehydrocholesterol to vit D3 (cholecalciferol). in skin. Vit D3 is than carried to the liver where it convert to 25 Hydroxycholecalciferol (25OHD3) which further convert into 1,25(OH)2D3 in kidney by enz (PTH stimulate this conversion). Vitamin D deficiency 1. Rickets (in children) -- soft bone, stunted growth, improper mineralization of bones. 2. Osteomalacia (in adults) -- Demineralization of existing bones, pathologic fracture. Bone demineralization may also occur due to Glucocorticoids which stimulate conversion of active Vit D intoInactive form.Vitamin D Toxicity: hypercalcemia, metastatic calcification, hypercalciuria, kidney stones, bone demineralization. * Vitamin E (antioxidant) protect mem & protein from free radical damage. It is present in green leafy vegi & seeds. Vit E also called Tocopherol react with free radical & form Tocopherol radical than Vit C than reduces the tocopherol radical and regenerate the tocopherol. Vitamin E deficiencyAtresia with impaired reflexes, myopathy with creatinuria, muscle weakness, and hemolytic anemia, retinal degeneration. Vit E + cause by malabsorption from cystic fibrosis, celiac dis, chronic cholestasis, pancreatic insufficiency,abetalipoproteinemia Vitamin E toxicity hypercoagulatory state * Vitamin K is required for post translational carboxylation of glutamyl residue in a no of Ca++ binding proteinnotably Clotting Factor VII, IX, X. Vit K is also produce by intestinal flora.Vitamin K deficiency results due to fat malabsorption, Drugs, Antibiotic (suppress flora). impaired blood clotting Vitamin K cross placenta poorly thats why single injection of vit K (0.5 - 1 mg) usually given to new born. Highdoses of Vit K can cause anemia, hyperbilirubiemia, kernicterus (accumulation of bilirubin in tissue). ------------------------------------------------------------------------------------------ * Ca++ :-is essential for bone, teeth, normal nerve and muscle function and normal clotting.Ca++ Def Paresthesia,| neuromuscular excitability, cramp, tetany, fracture, height loss, Osteomalacia & rickets Hypercalcemia caused by hyperparathyroidism, hypervitaminosis D, malignancy, hyperthyroidism (high boneturn over), Renal failure & drugs like lithium, thiazides, aluminum toxicity, milk alkali $. Some time it is idiopathic. * Iodine :- is essential for incorporation into thyroid hormones called organification. Iodine deficiency Goiter (enlarge thyroid gland), Cretinism (retarded growth and mental development in kids) | Iodine intake may also cause Goiter by blocking Organification. * Iron Present in Heme transport O2 (hemoglobin and Myoglobin). electron transport (cytochrome). Activation ofO2 (oxidases & oxygenases) Heme Iron absorbed by intestine more easily than Non Heme Iron . Ascorbic acid enhance Iron absorption where as antiacid and some plant food reduced Iron absorption. Iron deficiency cause 1. Hypochromic microcytic anemia. 2. Fatigue, Pallor, tachycardia, Dyspnea (shortness of breath) on exertion. 3. Burning sensation with depapillation of tongue. Iron toxicity cause Hemochromatosis and Hemosidrosis. KICK THE BOARDS. USMLE STEP 1 BIOCHEMISTRYPrepared by Dr. IRFAN MIR ONLY FOR EDUCATIONAL PURPOSE.14 NOT FOR COMMERCIAL USE OR SALE * Mg++ :- bind to active site of many enz and ATP. Mg++ deficiency seen in alcoholics, fat malabsorption & other malabsorption $ Neuromuscular excitability,muscle spasm, paresthesia. if deficiency prolongs tetany, seizure and coma occur. Severe hypomagnesemiasuppression of PTH release with resultant hypocalcaemia. vs.Mild Hypomagnesemiastimulates PTH secretion which | serum Ca++ by absorpton. * Phosphorus :- 85% of phosphorus present in bone minerals as a Ca phosphate. phosphate serve as blood buffer.phosphate esters are constituents of RNA and DNA, and phospholipids are major constituents of cell membrane. Phosphorus deficiency is usually a consequences of abnormal kidney function results in defective bone mineralization, retarded growth, skeletal deformity, and bone pain. phosphorus deficiency can also result into less O2 release from hemoglobin due to + RBC 2,3 biphosphoglycerate hypoxia. * Zinc :- essential for metallo enzymes. Zinc deficiency growth retardation and hypogonadism. Impair taste ,smell and poor appetite . Reduced immune function, mental lethargy, dry scaly skin. Zinc toxicity Vomiting diarrhea (by galvanized container), Neural damage (metal fume fever, zinc shakes)due to toxic fume inhalation. ---------------------------------------------------------------------- GENE EXPRESSION * Transcription occur from DNA to RNA (in nucleus) where as Translation is RNA to Protein (in cytoplasm). * Codons do not over lap and they are not separated by spacers so is CONTIGOUS. * There is more than one codon for some AA so it is DEGENERATE. * Each codon specifies only one AA so it is called UNAMBIGUOUS. * DNA contain transcription promoters, binding site for regulatory protein, and signal for gene rearrangement. * mRNA contain transcription terminator, processing signals, translation alignment signal, and start & stop codon. * In Eukaryotic cell replication and transcription occur in nucleus, where as translation occur in cytosol. * DNA is double stranded helix with strands anti parallel and complementary. * Anti parallel means one chain runs 5' 3' direction and other runs 3' 5' direction. * Complementary means that Adenine (A) always pair with Thymine (T) & Guanine (G) always pair with Cytosine (C) * DNA double stranded helix is stabilize by Hydrogen Bonds ( AT has 2 and GC has 3 H+ bonds ). * RNA contain uracil Instead of thymine as well as A, G, C. * RNA has single strand helix it may form internal double strand region called HAIR PIN LOOP. * Complimentary DNA and RNA strand can also associate or hybridize by denaturation. The presence of DNA, RNA known sequence may be detected by using hybridization probe. * DNA replication is semi conservative which means each daughter DNA contain one strand of parent DNA andone newly synthesized daughter DNA. * DNA polymerase catalyze DNA syn. DNA polymerase lll involve in replication. * Helicase ( DNA unwinding protein ) unwind the DNA duplex and Topoisomerase relieve the strain impose byunwinding ( unwinding occur by DNAgyrase in E. Coli ).* Primase make RNA primer to start DNA replication, RNA primer is complementary to DNA Templet and move in 5' 3' direction. * DNA Polymerase add nucleotide to the 3' OH end so called Leading strand grows continuously in 5' 3'direction. * The DNA Polymerase (DNA P) add incoming nucleotide only to 3' OH end, make DNA replication discontinuousin the other daughter strand called Lagging strand.DNA P lll built a DNA strand in the primer until it encounter the 5' end of the next Okazaki fragment. * New synthesized DNA segment called Okazaki fragment they are joined together to form continuous DNA chainby DNA ligase. KICK THE BOARDS. USMLE STEP 1 BIOCHEMISTRYPrepared by Dr. IRFAN MIR ONLY FOR EDUCATIONAL PURPOSE.15 NOT FOR COMMERCIAL USE OR SALE * DNA P also associated with 3' 5' Exonuclease activity which allows detection and removal of mismatch basepair. This correction is called EDITING. * When UV specific Endonuclease nicks one strand of the double stranded DNA, the DNA P synthesize new DNAstrand. Exonuclease activity of DNA P carries out 5' 3' excision of damage strand and finally DNA Ligase joinsthe ends together to complete repair. * Defective DNA repair cause Xeroderma pigmentosum, Fanconi $, Bloom $, etc. * RNA Polymerase (RNA P) catalyze Transcription. 1. RNAP I synthesize rRNA found in nucleolus and is resistant to inhibition by o- amantin (poison mushroom). 2. RNAP II synthesize mRNA found in nucleoplasm and inhibit by o- amantin 3. RNAP III synthesize tRNA and 5SrRNA found in nucleoplasm and moderately inhibit by o- amantin. * Mitochondrial RNAP transcribes RNA from mt gene. Mitochondrial RNAP inhibit by Refampicin but not o- amantin. * CAP at 5' end of the mRNA in eukaryotes protect against nuclease digestion & help align the mRNA properlyduring translation. * Poly - A - Tail in mRNA add to a 3' end. * mRNA primary transcript contain untranslated region called INTERVENING SEQUENCE or INTRON. Removal ofinteron called RNA splicing. * Intron begins with G-U and ends with A-G, small nuclear RNA assist the enz in order to make precise cut. In thecut region (intron) form LARIATSTRUCTURE which than removed and discarded. Later protein syn occur. * The enz Aminoacyl - tRNA synthatase form aminoacyl tRNAs (AA-tRNA) to start protein synthesis which is done bymRNA.* AA-tRNA binds to 60S and mRNA binds to 40S. * Translation (initiation) starts from start codon AUG at 3' end of the mRNA cap.(Remember AUG specifies Methionine). * Termination occur when ribosome encounters a non sense codon (termination) eg UAA, UAG, UGA which signaltermination. * Wobble is the codon in the mRNA (3 base) and anti codon on tRNA (5 base) it can wobble at the nucleotide nucleotide pairing site. [ these bases also called wobble base] ---------------------------------------------------------------------------- * Selective precipitation involve use of PH, heat, salt to separate protein from solution. * Gel Filtration and Polyacrylamide Gel Electrophoresis which separate protein on the bases of size. * Gel Electrophoresis and Ion Exchange Chromatography which separate protein on the bases of ionic change. * Affinity Chromatography involve removal of protein from mixture of specific binding on there ligands or to antibodies. * Edman Degradation Methodused to determine the AA sequence (primary structure). * Cyanogen Bromide cleaves peptide bond on the carboxyl side of methionine residue. * 2 - nitro - 5 - thiocyanobenzene cleave peptide bond on the AA side of cysteine residues. * Trypsin cleave peptide bond on the carboxyl side of lysine or arginine residues. * Chymotrypsin cleaves peptide bond on the carboxyl side of Aromatic and bulky non polar residue. * Gel Electrophoresis is used to separate DNA fragment on the basis of size. * Northern Blotting use hybridization probe to detect RNA fragment other wise this procedure is performed likesouthern blotting. * Southern blotting is used to detect DNA fragment that contain specific base sequence using auto radiography orFluorescence Southern blotting can also be used to visualized RFLPs. * Western blot use antibodies to detect protein. * Sanger Dideoxynucleotide Method and Maxam Gilbert Method are two techniques used to determine thesequences of the bases in DNA fragment. * PCR is used to amplify very tiny pieces of DNA. * ELISA is a rapid lab test in which Ab and Ag from pt is exposed Ag or Ab linked to an enz. A+ test result is Ag Ab match and indicate color change. KICK THE BOARDS. USMLE STEP 1 BIOCHEMISTRYPrepared by Dr. IRFAN MIR ONLY FOR EDUCATIONAL PURPOSE.16 NOT FOR COMMERCIAL USE OR SALE * DNA Finger Printing can be used to identify an individual DNA contain sequence variation called Polymorphism. * Some sequence polymorphism occur in or near the site of cutting by restriction enz. This lead to restrictionfragment length polymorphism (RFLP) which are different in size among individuals. ---------------------------------------------------------------------------- HORMONES * HORMONES1. Water soluble --- Catecholamine (eg EN) Peptide hormone (eg TRH) Protein hormone (eg Insulin) 2. Lipid soluble ----Steroid hormone Thyroid hormone* Water soluble hormones are free in blood where as lipid soluble hormones are bound to transport protein. * Water soluble hormones has mem receptorwhere as lipid soluble hormones has IC receptor. * Water Soluble Hormones :-1. Hormones bind to one group of receptors stimulate adenylate cyclase which convert ATP to cAMP than cAMPactivate protein kinase A which further phosphorylate several proteins. 2. Hormones binds to second group of receptors activate phospholipase C which hydrolyses phosphatidylinositol4,5biphosphate (PIP2) to yield IP3 & DAG, than DAG activates protein kinase A & IP3 stimulates the release ofCa++from ER. 3. Hormones bind to third group of receptors that stimulate Tyrosine Kinase activity which involveautophosphorylationof some of the receptors own Tyrosine residue. The phosphorylated receptors thaninteract with IC protein to alter cell activity. * Lipid Soluble Hormones pass through the cell mem and bind the IC hormone cell receptor protein to activate itthan activated receptor becomes transcription factor which bind to hormone response element in DNA andalter gene expression. * HORMONAL REGULATION OF FUEL METABOLISM :- 1. Insulin (secreted by | cell) lowers blood sugar & non esterified fatty acid by | glucose entry into the tissue , | glucose metabolism, | AA acid entry into muscle , + lipolysis and fatty acid release in adipose tissue . Hyperglycemia and AA stimulates insulin secretion where as EN + it. 2. Glucagon secreted by o cells in response to + blood glucose level (hypoglycemia) and | level of blood AA.*Glucagon stimulate Adenylate cyclase to production cAMP and activate protein kinase A.*Glucagon | glycogenolysis, | lipolysis, and fatty acid release in adipose tissue *Glucagon also | functioning of glucose - alanine cycle b/w liver and muscle. 3. Epinephrine (by adrenal medulla) elevate blood glucose & fatty acid (provoke the flight flee reflex) in responseto neural factor, hypoglycemia & hypoxia, by stimulating adenylate cyclase & resultant cAMP production &activation of protein kinase A. * EN | glycogenolysis, | lipolysis and fatty acid release in adipose tissue. | HR(chronotropic effect) & force of contraction (inotropic effect) & dilatation of blood vessels in muscle &constriction in skin & splanchnic blood vessels. 4. Cortisol (glucocorticoids) is secreted in response to ACTH (ACTH release by CRH from hypothalamus). | plasma Cortisol Reduced CRH secretion.Function of cortisol :- | Blood glucose. Inhibit insulin secretion & inflammation and Suppress immune system. | break down of muscle protein (release AA for gluconeogenesis) & | total body fat (at the| syn of gluconeogenic enzexpense of muscle protein) | water excretion by kidney It also | resistance to stress KICK THE BOARDS. USMLE STEP 1 BIOCHEMISTRYPrepared by Dr. IRFAN MIR ONLY FOR EDUCATIONAL PURPOSE.17 NOT FOR COMMERCIAL USE OR SALE HORMONAL REGULATION OF Na+ AND H2O BALANCE :- 1. Aldosterone secrete in response to renin angiotensin and | bloodK+ . It stimulate Na+ retention and K+secretion by kidney, Salivary gland and intestinal mucosa. 2. Arginine Vasopressin (ADH) is secreted by post pituitary in response to | plasma osmolality and neural impulses. It cause H2O reabsorption by kidney. -------------------------------------------------------------------------------------------------------- HORMONAL REGULATION OFCa++&PO4 METABOLISM :- 1. PTH is a protein secreted in response to hypocalcaemia to raise plasma Ca++ and lower plasma PO4. Where as hypercalcemia inhibit PTH secretion.PTH also stimulate osteoclast activity of bone to | blood Ca++and PO4. PTH + Ca++ excretion and | PO4 excretion by kidney. PTH also stimulate Calcitriol ( vit D) formation by kidney with resultant | absorption of Ca++by intestine. 2. Calcitriol ( 1,25 (OH)2 - D3 ) is derived from vit D3. Vit D3 syn involve 2 hydroxylation reaction 1st in liver and 2nd in kidney. Hypocalcaemia and PTH stimulates second hydroxylation reaction in kidneyproduce CalcitriolCalcitriol stimulate Ca++absorption from gut and | efficiency of PTH on bones (osteoclastic activity) |plasma Ca++ &PO4. . ------------------------------------------------------------------------------------------------------ HORMONAL REGULATION OF BODY METABOLISM :- 1. Thyroxine and Triidothyronine secreted by Thyroid follicle cells in response to TSH from Ant pituitary.TSH secretion is stimulated by TRH, | level of plasma Thyroxine inhibit TRH secretion. Thyroxine and Triidothyronine| the metabolic rate through out body. It cause | heat production, | growth, |mental activity, | sensitivity of tissue to EN, | catabolism of cholesterol + blood cholesterol. 2. Human Growth hormon (HGH) is a protein secreted from Ant pituitary in response to GHRH.GHRH inhibit by Somatostatin and GH releasing inhibitory hormone from Hypothalamus.GH stimulate liver to secrete Insulin like Growth Factor I which has anabolic effect by increasing protein syn, |bone calcification and matrix formation, and | AA uptake by muscle, bone and kidney. GH also | blood glucose (insulin antagonist)and | fatty acid release from adipose tissue. ----------------------------------------------------------------------------------------- HORMONAL REGULATION OF REPRODUCTIVE SYSTEM :- 1. FSH and LH : GnRH (from hypothalamus) which regulates FSH and LH secretion. 1. Testosterone --- LH stimulate Leydig cells to synthesize and secrete testosteron. whereas FSH along with testosterone stimulate spermatogenesis in sertoli cells. Primary Function of Testosterone :- Spermatogenesis (in conjunction with FSH), Maturation, function of prostate &seminal vesicle, Maturation of male sex organ (penis and scrotum)Interest & ability to engage in sexual activity Other function of testosterone :- Pubertal growth spurt, Maturation of skin Male pattern of hair distribution, Deepening of voice & aggressive personality. 2. Estradiol is phenolic steroid secreted by ovarian follicles. It prepare female reproductive sys for pregnancy. Function of Estradiol :- Maturation of uterus cervix and vagina Proliferation of vagianl epithelium and uterine endothelium Duct proliferation and fat deposition in mammary gland. Other function of Estradiol :- | bone calcification and closure of epiphysis. Skin maturation and female pattern of hair and fat distribution.KICK THE BOARDS. USMLE STEP 1 BIOCHEMISTRYPrepared by Dr. IRFAN MIR ONLY FOR EDUCATIONAL PURPOSE.18 NOT FOR COMMERCIAL USE OR SALE 3. Progesterone is secreted by Corpus luteum and promote gastation in women by 1. Transformation of proliferative endometrium to seceretory endometrium. 2. Prevention of synchronized uterine muscle contraction. 3. maintainance of pregnancy 4. It also stimulate growth of mammary gland for milk secretion 4. GnRH release in cyclic secretion (responsible fo menstrual cycle) and result in FSH and LH secretion 1. FSH stimulate growth of ovarian follicles and secretion of estradiol by follicular Granulosa cell. 2. A burst of LH and FSH secretion stimulate ovulation by rupturing the follicles. 3. LH stimulate formation and function of Corpus luteum which secrete Estradiol and Progesterone 5. PL secrete from Ant pituitary stimulate milk production in mammary gland that have been prepared by estradiol& progesterone. 6. Oxitocin form Post pituitary stimulate milk ejection and uterine contraction. ------------------------------------------------------------------------------ KEY POINTS TO REMEMBER * Key Differences :- 1. Purine A - G have two rings. 2. Pyrimidine CTU have one ring. 3. Guanine has ketone. 4. Thymine has methyl. * DNA repair enz :- 1. Specific Glycosylase Recognize and remove damaged DNA. 2. Endonuclease 3. DNA polymerase 4. DNA ligase. * Exonuclease remove short stretch of nucleotide . * DNA and RNA synthesize in 5'3' direction (remember 5' end bears triphosphate which is energy source). * Protein synthesize in 5'3' direction . * Primase makes RNA primer on which DNA polymerase can initiate replication. * mRNA is largest RNA in size where as tRNA is smallest in size. * rRNA is most abundant. * Creatinine or urea both made from arginine. * Glycine is use to make porphyrin which is than used to make heme. * No male to male transmission is characteristic of X - Linked Ressesive Disorder. * Most distinguihing finding in Tay- sachs is cherry red macula.