the chemical level of organization 25 · 2-1 introduction – chemical level of organization •...

© 2012 Pearson Education, Inc.

PowerPoint® Lecture Presentations prepared by Jason LaPres Lone Star College—North Harris

2 The Chemical Level of Organization

+ 25 Basics of Metabolism & Energetics

NOTE: Presentations extensively modi6ied for use in MCB 244 & 246 at the University of Illinois by Drs. Kwast & Brown (2013-‐2014)


•  Distinguish among the types of chemical reactions that are important in physiology (Chapter 2).

•  Understand fundamental thermodynamics of chemical reactions and the role of enzymes in metabolism (Chapter 2).

•  Have a general understanding of—and conceptual appreciation for—the overall design of cellular metabolism (Chapter 25, pp. 916 – 934 [sections 1 - 4]).

Chapters 2 + 25 Learning Objectives

2


2-1 Introduction to Chemical Level of Organization

•  Prerequisite for MCB 244 is CHEM 101 & 102

•  Thus, I will assume you have a working

knowledge of basic chemistry including:

•  Atoms and subatomic particles structure/function…

•  Nature of various chemical bonds and reactions

•  Basic energy concepts

If you do not have a working knowledge of these

subjects, read all of chapter 2 as a review. 3


2-1 Introduction – Chemical Level of Organization •  Chemical bonds involve the sharing, gaining, and

losing electrons in the valence shell •  Three majors types of chemical bonds:

•  Ionic bonds: •  attraction between cations (electron donor) and anions (electron

acceptor)

•  Covalent bonds: •  strong electron bonds involving shared electrons

•  Hydrogen bonds: •  weak polar bonds based on partial electrical attractions

•  In addition other important interactions include Van der Waals attractions, hydrophobic forces and electrostatic attractions

•  As we shall see, much of the chemistry of life involves weak, non-covalent interactions, especially for proper enzyme functioning 4


2-1 Atoms and Atomic Structure

•  Basic Energy Concepts – Read the book if you are unfamiliar with these concepts:

•  Energy •  The power to do work

•  Work •  A change in mass or distance

•  Kinetic energy •  Energy of motion

•  Potential energy •  Stored energy (e.g., chemical, mechanical, electrical)

5


2-1 Introduction – Chemical Level of Organization •  Prerequisite for MCB 244 is CHEM 101 & 102

•  Thus, you are expected to have a working knowledge of basic

chemistry including:

•  Atoms and subatomic particle structure/function

•  Chemical bonds and reactions

•  Basic energy concepts

•  We’ll begin formal lecture with basic enzymology in Chapter 2, p. 37

(see pp. 26 – 37 if you need a refresher on the above).

•  Please note that additional information is provided herein that

you are responsible for but is not contained in your text (e.g.,

basic metabolism, enzymology and thermodynamics.) 6


2-4 Basics of Metabolism and Enzymes

•  The chemistry of life is organized into metabolic

pathways NOTE: Parts of this section are from pp. 916 – 926 (Chpt 25) BUT you

are only responsible for the general overview I present here in slide and verbal form.

•  The totality of an organism’s chemical reactions is called

metabolism.

•  Furthermore, these reactions can be divided into those that break

down molecules into smaller units (catabolism) versus those that

build up molecules into large ones (anabolism).

•  A cell’s metabolism is an elaborate road map of the chemical

reactions in that cell.

•  Metabolic pathways alter molecules in a series of steps. 7



•  Pictured at the right is a ball

and stick ball diagram of

pathways involved in basic

cellular metabolism, where

balls represent molecules

and sticks interconnections.

•  Can you name the

metabolic pathways

pictured in bold? 8


There are “Quite a Few” Pathways in Metabolism

Map 1 of 2 of the basic metabolic pathways

9



•  Enzymes selectively accelerate each step.

•  The activity of enzymes is regulated to maintain an appropriate balance of

supply and demand.

•  Catabolic pathways release energy by breaking down complex

molecules to simpler compounds.

•  This energy is stored in organic molecules until need to do work in the cell.

•  Anabolic pathways consume energy to build complicated

molecules from simpler compounds.

•  The energy released by catabolic pathways is used to drive

anabolic pathways.

10



•  The concept of free energy provides a criterion for measuring the spontaneity of a system.

•  Free energy is the portion of a system’s energy that is able to perform work (when temperature is uniform).

11


Chemical Reactions, Thermodynamics and Enzymes

Basic Thermodynamics: •  1st law - energy is neither created or destroyed (conservation of energy)

•  2nd law - entropy (S, measure of randomness) progressively increases in a closed system •  So what about highly ordered and structured biological systems?

•  Don’t they violate the 2nd law?

•  G (free energy) and entropy (S) are related by the following equation:

ΔG = ΔH – TΔS

where ΔH is the change in heat or enthalpy (ΔH) and T is the absolute temperature (T)

Given that ΔS tends to increase and the fact that living system operate within very narrow ranges of temperature (hence ΔH ≈ 0), chemical reactions that result in a reduction in free energy (ΔG < 0) will occur “spontaneously”.

•  Let’s look next at how this relates to chemical reactions… 12


Chemical Reactions, Thermodynamics and Enzymes A + B ↔ C + D

(as written “A” and “B” are reactants and “C” and “D” are products) •  K’eq = [C] [D] / [A] [B]

•  ΔGº = -RT ln(K’eq)

•  If K’eq > 1, then ΔGº is negative [exergonic rxn] and the reaction will spontaneously proceed in the forward direction as written (NOTE: activation energy [ΔG*] must still be overcome—see following slides)

•  If K’eq < 1, then ΔGº is positive [endergonic rxn]. Energy input required for the reaction to go in the forward direction (e.g., transfer of chemical energy by coupled reaction [ATP → ADP, -7.3 kcal/mol])

•  Let’s now turn to the role of enzymes and their effect on ΔG*

13


2-4 Enzymes are catalysts that lower the activation energy (ΔG*) •  Your book says ”Chemical reactions in cells cannot start without

help.” In reality, the probability of some reactions happening is infinitesimally small and obtaining equilibrium exceedingly slow (think geological time frames) because of high activation energy.

•  Activation energy (ΔG*) is the minimum amount of energy needed to get a chemical reaction “started”.

•  Enzymes are catalysts, typically proteins, that lower the activation energy of reactions.

•  Catalytic power comes from their ability to bind substrate molecule(s) in precise orientations and stabilize transition states in the making and breaking of chemical bonds

•  Enzymes typically speed up reaction rates by 108 – 1020 times!

•  Enzymes are not consumed by the reactions they catalyze, nor do they alter the equilibrium of these reactions

•  Enzymes catalyze some 4,000 biochemical reactions in our bodies

14


Enzyme Types and Reactions Rates

15


2-4 Enzymes lower ∆G*

E + S

E = Enzyme

S = Substrate

P = Product

ES*

E + P

ΔG◦

ΔG*

ΔG*

Figure 2-8 ( p. 37)

16


2-11 Enzyme Function •  Enzymes are very specific in terms of what they bind. The substance an

enzyme acts upon (usually by binding) is called a substrate.

•  The active site of an enzymes is typically a pocket or groove on the surface of the protein into which the substrate fits.

•  The specificity of an enzyme is due to the fit between the active site and that of the substrate.

•  In most cases substrates are held in the active site by weak interactions, such as hydrogen bonds and ionic bonds.

Substrates bind to active site of enzyme

ENZYME Active site

Substrates S1

S2

Once bound to the active site, the substrates are held together and their interaction facilitated

ENZYME

S1 S2

Enzyme-substrate complex

Substrate binding alters the shape of the enzyme, and this change promotes product formation

PRODUCT

ENZYME

Product detaches from enzyme; entire process can now be repeated

ENZYME

FIGURE 2–22 A Simplified View of Enzyme Structure and Function.

17


2-11 Modifications to Enzyme Function

•  Binding by some molecules (e.g., inhibitors) prevent enzymes from catalyzing reactions.

•  If the inhibitor binds to the same site as the substrate (the active site), then it blocks substrate binding via COMPETITIVE inhibition.

•  NONCOMPETITIVE inhibitors do not interact with the enzyme’s active site but rather bind to another part of an enzyme molecule (allosteric site) thereby modulating function.

18


2-11 Cofactors, Coenzymes & Enzyme Variants Cofactors

•  An ion (e.g., Zn, Mg, Fe, etc.) or organic molecule (e.g., heme) that binds to an enzyme thereby enabling the “active site” to bind substrate(s)

“Coenzymes” •  Are small organic molecules that transport chemical groups from one

enzyme to another (e.g., NAD+/NADH, NADP+/NADPH, FAD+/FADH2 etc. )

Isozyme •  Different form of an enzyme that catalyzes the same reaction but is

encoded for by a different genetic locus (i.e., they are different genes; e.g., tissue-specific isozymes)

Allozyme •  Different form of an enzyme that catalyzes the same reaction and is

encoded by the same genetic locus (i.e., they are encoded by the same gene but their DNA and amino acid sequence are different. NOTE: Not possible in haploid state)

19


Example of Tissue-Specific Isozymes - LDH Lactate dehydrogenase catalyzes the interconversion of pyruvate and lactate with concomitant interconversion of NADH and NAD+.

Functional lactate dehydrogenases come in 2 different “flavors”: homo-tetramers (4 identical subunits) and heterotetramers composed of M- and H-type subunits that are encoded for by the LDHA and LDHB genes, respectively:

* LDH-1 (4H) - in the heart and RBCs

* LDH-2 (3H1M) - in the reticuloendothelial system

* LDH-3 (2H2M) - in the lungs

* LDH-4 (1H3M) - in the kidneys, placenta and pancreas

* LDH-5 (4M) - in skeletal muscle and liver 20


Basic Enzyme Kinetics

P EES S E 31

2

kk

k+⎯→⎯+ ↔

E = Enzyme S = Substrate P = Product

ES = Enzyme-Substrate complex

k1 = rate constant for the forward reaction

k2 = rate constant for the breakdown of the ES to substrate (reverse rxn to k1

k3 = rate constant for the formation of the products 21


Michaelis-Menten Kinetics

V0 = Vmax • [S] ! [S] + Km!

E + S⇔ES⇒E + P!

Michaelis-Menten Model!

Km = [S] at ½ Vmax Km =provides a relative measure (inverse) of the affinity of an enzyme for its substrate; lower Km = higher affinity and thus higher velocity V0 (initial velocity)

= moles product formed per sec. 22


Figure 25-1 An Introduction to Cellular Metabolism INTERSTITIAL

FLUID

Organic Molecules • Amino acids • Lipids • Simple sugars

CATABOLISM

NUTRIENT POOL

HEAT

ANABOLISM

Plasma membrane

Results of Anabolism

• Maintenance and repairs • Growth • Secretion • Stored nutrient reserves

Anaerobic catabolism in the cytosol releases small amounts of ATP that are significant only under unusual conditions.

Aerobic Metabolism (in mitochondria)

CYTOPLASM

60% 40%

Other ATP EXpenses • Locomotion • Contraction • Intracellular transport • Cytokinesis • Endocytosis • Exocytosis

ATP

ATP

23


Organic compounds that can be absorbed by cells are distributed to cells throughout the body by the bloodstream.

KEY

MITOCHONDRIA

Nutrient pool

= Catabolic pathway

= Anabolic pathway

Three-carbon chains

Two-carbon chains

Fatty acids Glucose Amino acids

Triglycerides Glycogen Proteins

Citric acid cycle

Coenzymes Electron transport system

Structural, functional, and storage components Figure 25-2 Nutrient Use in Cellular Metabolism

24

*Catabolism Overview*

Lipids (fats - e.g., triglyceride) --broken down into fatty acids (FAs) and glycerol in cytoplasm; FA → acylCoA and enters mitochondria --acylCoA shortened 2 carbons at a time during conversion to acetyl-CoA --acetyl-CoA enters Krebs in mitochondria Polysaccharides (carbohydrates -e.g., glycogen) --broken down to constitutive hexose (e.g., glucose) or pentose sugars --glucose broken down to pyruvate via glycolysis in the cytoplasm with net yield of 2 ATP --when O2 is limiting, pyruvate converted to lactate to preserve cellular redox balance (NAD+) --pyruvate enters Krebs as acetyl-CoA

Proteins (NOT stored for catabolic purposes) --broken down into constitutive amino acids --amino acids degraded via trans- or de-amination -- and carbon atoms fed into the Kreb’s cycle at various points

mitochondria Acetyl CoA

Electron Transport

Chain ATP

Fatty Acids, Glycerol

Pyruvate

Proteins Polysaccharides

O2

Lipids

5- & 6-carbon sugars

3-carbon sugars

ATP

H2O

Lactate

CO2 TCA or Krebs Cycle

Res

pira

tion

(aer

obic

)

Gly

coly

sis

(ana

erob

ic)

H+

Amino Acids

cytoplasm


25-3 Glycolysis THE BASICS

•  Location: cytoplasm

(exception: sperm LDH)

•  Glucose → pyruvate or lactate

•  One oxidation step balanced by one reduction step IF lactate formed

•  ATP required at 2 steps

•  ATP generated at 2 steps after conversion to 3-C sugar for net yield of 2 ATP

•  Does not require O2

•  ATP required at 2 steps

•  High power output (nearly 2x that of glucose oxidation)

OR-- lactate

lactate NADH

NAD+

26


25-4 The Citric Acid Cycle THE BASICS

• Location: mitochondrial matrix

• Catabolic Function: Remove H+s from acetyl-CoA and transfer to “coenzymes” (NADH & FADH2)

• 1 GTP formed per cycle (0 ATP)—2 cycles per acetyl-CoA

• 3 NADH and 1 FADH2 formed per cycle

• Referred to as “hub of metabolism” as final pathway for complete oxidation of fats, carbohydrates & proteins

• O2 not required per se

• Amphibolic Pathway (serves 2 purposes): oxidation of acetyl-CoA and precursors for bio-synthesis (green arrows)

• 

Aspartate Synthesis

Fatty Acid Synthesis

Heme Synthesis

Glutamate Synthesis

27


25-5b Oxidative Phosphorylation

I

II

III IV V

ATP

ADP/ATP-Translocase

H+ Pi

H+/Pi Symport

THE BASICS

• Location: Inner mitochondrial membrane

• Electrons from NADH & FADH2 + O2 → H2O

• As electrons passed down, free energy loss coupled to proton (H+) pumping (matrix → intermembrane space)

• Complexes I,III & IV pump protons

• Displacement of H+s creates both chemical (ΔpH) and electrical gradient (ΔΨ) = proton motive force

• ATP-synthase uses this force to couple movement of protons back into matrix with phosphorylation of ADP to ATP 28


25-6 Energy Yield & Substrate Comparison

020406080100

FA C6-Ferm.

PowerAvailability

Power Output vs. Energy Availability (cyan) (indigo)

Power output = µmol ATP • g-1 •min-1

Availability = µmol ~P • g wet weight-1

•  Actual yield is closer to 25.2 mol of ATP/mol glucose, NOT 36.

•  WHY? Cost of transporting ATP-4 out of matrix and ADP-3 into matrix and exchange of Pi for H+ (i.e., these processes degrade the proton gradient)

Substrate Comparison Per gram, fats yield about 2.3x more ATP than carbo’s or aa’s—Why?—they are more reduced thus yielding more NADH & FADH2 per gram, however…

29


2-25 ATP Structure/Function THE BASICS • Ubiquitous role in energy metabolism, biosynthesis, contractions, ion pumping, etc. (i.e., in work) • Hydrolysis of a phosphate group forms adenosine diphosphate [ATP -> ADP + Pi] and releases 7.3 kcal of energy per mole of ATP under standard conditions.

• Although the phosphate bonds of ATP are sometimes referred to as “high-energy bonds”, they are actually fairly weak covalent bonds that are unstable because of the negative charge of each phosphate group. • The energy from the hydrolysis of ATP is coupled directly to endergonic processes by transferring the phosphate group to another molecule, thereby driving the endergonic reaction. • Often we refer to energy conversions in ATP equivalents: NADH → NAD+ = 3 ATPs iff NADH in mito matrix FADH2 → FAD+ = 2 ATP equivalents NADPH → NADP+ = 4 ATP equivalents

30


Redox Reactions, Reducing Power and the different cellular roles of NADH & NADPH

•  Redox (reduction-oxidation reaction) describes all chemical reactions in which atoms have their oxidation number (oxidation state) changed.

•  Oxidation is the loss of electrons or an increase in oxidation state whereas

•  Reduction is the gain of electrons or a decrease in oxidation state.

•  The main function of the “coenzyme” NAD+ is as a oxidizing agent (electron acceptor) in cellular catabolism (e.g., TCA), thus the NAD+/NADH ratio is kept very high in the cell (strong oxidizing potential).

•  The main function of the “coenzyme” NADPH is as a reducing agent (electron donor) in anabolism, thus the NADP+/NADPH ratio is kept very low in the cell (strong reducing potential).

31


Major Functional Units in Cellular Metabolism •  Simplified engineering diagram of the major functional blocks in cellular metabolism

(after Hochachka & Somero: Biochemical Adaptation) •  3 Fundamental Requirements of All Cells: Energy (ATP), Reducing Equivalents

(NADPH) & Biosynthetic Precursors

Proteins Nucleic Acids Membranes Organelles

BLOCK I

CATABOLISM

BLOCK II

BIOSYNTHESIS

MECHANICAL WORK

CHEMICAL WORK

BLOCK III

GROWTH

&

INTEGRATION

NAD+

NADP+

NADH

NADPH

ATP

ADP

NDP

NTP ATP

ADP

Nutrients & Metabolites Glycolysis Pentose

Phosphate TCA

Amino Acids Nucleotides

Hormones, etc.

NTP = Nucleotide Triphosphate (e.g., GTP, UTP, CTP) 32


2-14 Recycling of Cellular Components

•  Colloids and Suspensions •  Colloid

•  A solution of very large organic molecules

•  For example, blood plasma

•  Suspension

•  A solution in which particles settle (sediment)

•  For example, whole blood

•  Concentration

•  The amount of solute in a solvent (mol/L, mg/mL)

33


Chapter 2+25 Summary Questions/Goals •  What is free energy and how is it used to perform useful work in cells?

•  What is an enzyme and what are their unique roles in cellular metabolism?

•  Understand the basics of enzyme catalyzed reactions and their kinetics as provided in class. (Be able to interpret graphs of velocity vs. substrate concentration.)

•  Describe the (a) overall function, (b) starting and ending materials as well as any important intermediates, and (c) the cellular compartment in which each of the following metabolic pathways occur:

Glycolysis Citric Acid or TCA or Kreb’s Cycle Electron Transport

•  Describe the unique roles that both ATP and NAD(P)H have in cellular metabolism.

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the chemical level of organization 25 · 2-1 introduction – chemical level of organization •...

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