protein structure cbmol4 5

8/14/2019 Protein Structure CBMol4 5

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Peptide bond

Amino acid

C-terminus

terminates by a

carboxyl group

N-terminus

terminatesby an amino group

A peptide: Phe-Ser-Glu-Lys (F-S-E-K)

From amino acids to protein:


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The Shape of proteins:

Occurs

Spontaneously

Native conformation

determined by

different Levels

of structure


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Tertiary structureoverall three-dimensional form of a polypeptide chain, which is stabilized by multiple

non-covalent interactions between side chains.

Primary structureThe linear arrangement (sequence) of amino acids and the location of covalent (mostly

disulfide) bonds within a polypeptide chain. Determined by the genetic code.

Secondary structure

local folding of a polypeptide chain into regular structures including the helix, sheet, and U-shaped turns and loops.

Quaternary structure:The number and relative positions of the polypeptide chains in multisubunit

proteins. Not all protein have a quaternary structure.

Four Levels of Structure Determine the

Shape of Proteins


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Bovine Insulin: the first sequenced protein

In 1953, Frederick Sanger determined the amino acid sequence of insulin, aprotein hormone .

This work is a landmark in biochemistry because it showed for the first time thata protein has a precisely defined amino acid sequence.

it demonstrated that insulin consists only of amino acids linked by peptide bondsbetween -amino and -carboxyl groups.

the complete amino acid sequences of more than 100,000 proteins are nowknown.

Each protein has a unique, precisely defined amino acid sequence.

Primary Structure of a protein:determined by the nucleotide sequence of its gene


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C-peptide

Hu

man: Thr-Ser-Ile

Cow: Ala-Ser-Val

Pig: Thr-Ser-Ile

Chiken: His-Asn-Thr

Pro-insulin protein

Insuline

+ C peptide

C-peptide

Pro-insulin is produced

in the Pancreatic islet

cells

Primary Structure

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Amino acid substitution in proteins from different species

Conservative Substitution of an amino acid by another

amino acid of similar polarity(Val for Ile in position 10 of insulin)

Non conservative

Substitution involving replacement

of an amino acid by another of

different polarity(sickle cell anemia, 6th position of hemoglobin

replace from a glutamic acid to a valine induceprecipitation of hemoglobin in red blood cells)

Invariant residues Amino acid found at the same position in

different species

(critical for for the sructure or function of the protein)


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SECONDARY STRUCTURE

Stabilized by hydrogen bonds

H- bonds are between CO and NHgroups of peptide backbone

H-bonds are either intra- or inter-

molecular

3 types : a-helix, b-sheet and triple-helix


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What forces determine the

structure?

Primary structure - determined by

covalentbonds

Secondary, Tertiary, Quaternary structures -

all determined by weak forces Weak forces - H-bonds, ionic interactions, van

der Waals interactions, hydrophobicinteractions


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Non covalent

interactions

involved in theshape of proteins


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Secondary structures:

Helix: helix conformation was discovered 50 years ago in keratine abundant in hair nails, and horns

Sheet:discovered within a year of the discovery of

helix.Found in protein fibroin the majorconstituant of silk


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The helix:result from hydrogen bonding, does not involve the side chain of the amino acid


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sheet:result from hydrogen bonding, does not involve the side chain of the amino acid


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Two type of Sheetstructures

An anti paralellel sheet

A paralellel sheet


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TRIPLE HELIX

Limited to tropocollagen molecule

Sequence motif of (Gly-X-Pro/Hypro)n-

3 left-handed helices wound together to give aright-handed superhelix

Stable superhelix : glycines located on thecentral axis (small R group) of triple helix

One interchain H-bond for each triplet of aas between NH of Gly and CO of X (or Proline) inthe adjacent chain


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Triple helix of Collagen


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NONREPETITIVE STRUCTURES

Helices/-sheets: ~50% of regular2ostructures of globular proteins

Remaining : coil or loop conformation Also quite regular, but difficult to

describe

Examples : reverse turns, -bends(connect successive strands ofantiparallel -sheets)


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The Beta Turn(aka beta bend, tight turn)

allows the peptide chain to reverse direction

carbonyl O of one residue is H-bonded to the amide proton of a residue

three residues away

proline and glycine are prevalent in beta turns (?)


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-bulge A strand of polypeptide in a -sheet may contain an extra residue

This extra residue is not hydrogen bonded to a neighbouring strand

This is known as a -bulge.


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The secondary structure of atelephone cord

A telephone cord, specifically the coilof a telephone cord, can be used asan analogy to the alpha helixsecondary structure of a protein.

The tertiary structure of a telephonecord

The tertiary structure of a protein refersto the way the secondary structure foldsback upon itself or twists around to forma three-dimensional structure. Thesecondary coil structure is still there, butthe tertiary tangle has beensuperimposed on it.

Tertiary structure: the overall shape of a protein

or a telephone cord!!!


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Tertiary structure: the overall shape of a protein

Full three dimensional organization of a protein

The three-dimensional

structure of a protein

kinase


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TERTIARY STRUCTURE

R-group interactions result in 3D structures ofglobular proteins

Types of interactions : H-, ionic- (salt linkage),

hydrophobic- and disulphide- bond Hydrophilic R groups on surface while

hydrophobic R groups buried inside of

molecule Wide variety of 3o structures: since large

variation in protein sizes and amino acidsequences


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The role of side chain in the

shape of proteinsWhere is water?

Hydrophobic

Hydrophilic


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Similarly:The tertiary structure for myoglobin is fairly well understood. Myoglobin has an alpha helix which then can be

viewed as being enclosed in this blue sheath, the sheath doesn't exist but we can draw it that way. That helix

folds back upon itself into what's referred to as the tertiary structure of myoglobin. Bonds between the side

groups of the amino acid residues are responsible for holding together the tertiary structure of this protein.


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A coiled-coil:Structure occurs when the 2 a helix

have most of their nonpolar

(hydrophobic) side chains on one

side, so that they can twist around

each other with these side chain

facing inwards


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Quaternery

structure:

If protein is formed as a

complex of more than one

protein chain, the complete

structure is designed as

quaternery structure:

Generally formed

by non-covalent

interactions between

subunits

Either as homo- or

hetero-multimers


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QUATERNARY STRUCTURE:

ADVANTAGES

Oligomers (multimers) are more stable than dissociated

subunits

They prolong life of protein in vivo

Active sites can be formed by residues from adjacent

subunits/chains

A subunit may not constitute a complete active site

Error of synthesis is greater for longer polypeptide chains Subunit interactions : cooperativity/ allosteric effects


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Primary structure

Secondary structure

Tertiary structure

Quaternary structure


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QuickTime and aTIFF (LZW) decompressor

are needed to see this picture.

Structure of the hemaglutinine

protein(a long multimeric molecule whose three identical subunits are each composed of twochains, HA1 and HA2).

Primary structure(1 letter code used)

Secondary structure

helicesstrandsrandom

coils


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QuickTime and aTIFF (LZW) decompressorare needed to see this picture.

Tertiary

structureQuaternary

structure

Protein

domains


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Protein domains:

Any part of a protein that can foldindependently into a compact, stable

structure. A domain usually contains

between 40 and 350 amino acids.

A domain is the modular unit from which

many larger proteins are constructed.

The different domain of protein are often

associated with different functions.


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The Src protein


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A X diff ti i f th t i l bi


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An X-ray diffraction image for the protein myoglobin.

The first protein crystal structure was of sperm whalemyoglobin, as determined by Max Perutz and Sir John Cowdery

Kendrew in 1958, which led to a Nobel Prize in Chemistry.

The X-ray diffraction analysis of myoglobin was originallymotivated by the observation of myoglobin crystals in driedpools of blood on the decks of whaling ships.


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NMR is a field of structural biology, that applies nuclearmagnetic resonance spectroscopy to investigating proteins

The field was pioneered by among others, Kurt Wthrich, who won the Nobel prize in 2002,

Pacific Northwest National Laboratory's highmagnetic field (800 MHz) NMR spectrometer being

loaded with a sample.

The NMR sample isprepared in a thin walled

glass tube.

Protein NMR is performed on aqueous samples of highly purified protein.

Sample consist of between 300 and 600 microlitres with a protein concentration in therange 0.1 3 millimoles.

The source of the protein can be either natural or produced in an expression system using

recombinant DNA techniques through genetic engineering.


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Function of

peptides and proteins


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Read Table 3.2 in Devlin

for other examples of

biologically active peptides

Oxytocin and vasopressin are two peptide hormoneswith very similar structure, but with very differentbiological activities.

Interestingly, their structures only differ by one aminoacid residue (the hydrophobic LEU number 8 in

oxytocin is replaced by a hydrophilic ARG residue invasopressin).

Oxytocin is a potent stimulator of uterine smoothmuscle, and also stimulates lactation.

Vasopressin, also know as antidiuretic hormone(ADH), has no effect on uterine smooth muscle, but

causes reabsorbtion of water by the kidney, thusincreasing blood pressure.


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Function of proteins

Proteins

are the most

important buffers in

the body.

Enzymatic catalysis

Transport and storage (the protein hemoglobin,albumins) Coordinated motion (actin and myosin).

Mechanical support (collagen).

Immune protection (antibodies)

Generation and transmission of nerve impulses

- some amino acids act as neurotransmitters,

receptors for neurotransmitters, drugs, etc. are

protein in nature. (the acetylcholine receptor),

Control of growth and differentiation -

transcription factors

Hormones

growth factors ( insulin or thyroid stimulating

hormone)

Why? Protein molecules

possess basic andacidic groups whichact as H+ acceptorsor donorsrespectively if H+ isadded or removed.


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Proteins are the most important buffers in the body. They aremainly intracellular and include haemoglobin.

The plasma proteins are buffers but the absolute amount issmall compared to intracellular protein.

Protein molecules possess basic and acidic groups which act asH+ acceptors or donors respectively if H+ is added or removed.

Many proteins (thousands!) present in blood plasma

Proteins contain weakly acidic (glutamate, aspartate) and basic(lysine, arginine, histidine) side chains (or R groups)

At neutral pH, only histidine residues (containing imidazole R

group with pKa ~ 6.0) in proteins can act as a buffer component

Haemoglobin with 38 histidine/tetramer is a good buffer

N-terminal groups of proteins (pKa ~ 8.0) can also act as a

buffer component


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Proteins play crucial roles in almost every biological process.They are responsible in one form or another for a variety ofphysiological functions including

Enzymatic catalysis

Transport and storage

Coordinated motion

Mechanical support

Immune protection

Generation and transmission of nerve impulsesControl of growth and differentiation


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Enzymatic catalysis -almost all biological reactions are enzyme catalyzed.

Enzymes are known to increase the rate of a biological reaction by a factor of 10 to the 6th power!

There are several thousand enzymes which have been identified to date.


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Transport and storage - small molecules are often carried by proteins in thephysiological setting (for example, the protein hemoglobin is responsible for the transport ofoxygen to tissues). Many drug molecules are partially bound to serum albumins in the plasma.

3-dimensional structure ofhemoglobin. The four subunits areshown in red and yellow, and theheme groups in green.

The binding of oxygen is affected by molecules such ascarbon monoxide (CO) (for example from tobacco smoking,cars and furnaces).

CO competes with oxygen at the heme binding site.Hemoglobin binding affinity for CO is 200 times greater thanits affinity for oxygen, meaning that small amounts of COdramatically reduces hemoglobin's ability to transportoxygen. When hemoglobin combines with CO, it forms avery bright red compound called carboxyhemoglobin.

When inspired air contains CO levels as low as 0.02%,headache and nausea occur; if the CO concentration isincreased to 0.1%, unconsciousness will follow. In heavysmokers, up to 20% of the oxygen-active sites can beblocked by CO.

C di t d ti


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Coordinated motion - muscle is mostly protein, and muscle contraction is mediated bythe sliding motion of two protein filaments, actin and myosin.

Platelet before activation Activated plateletActivated platelet

at a later stage than C)

Platelet activation is a controlled

sequence of actin filament:

Severing

Uncapping

Elongating

Cross linking

That creates a dramatic shape changein the platelet


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Immune protection - antibodies are protein structures that are responsible forreacting with specific foreign substances in the body.


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Control of growth and differentiation -

proteins can be critical to the control of growth, cell differentiation and expression of DNA.

For example, repressor proteins may bind to specific segments of DNA, preventing expression andthus the formation of the product of that DNA segment.

Also, many hormones and growth factors that regulate cell function, such as insulin or thyroidstimulating hormone are proteins.

Insuline


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Membrane transport proteins

STRUCTURE FUNCTION


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STRUCTURE - FUNCTION

RELATIONSHIPS

In general, all globular proteins have

distinctive

3D structures that are

specialized for their particular functions.

Sh d f i


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Shape and function

The relationship between shape and function of proteins:


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Protein degradation:


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Protein degradation:


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HOT Areas of Medical Research


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Human Genome

sequencing is completed

Application in Biology and

Medicine just beginning

e.g., Cloning of a disease gene is

the first step in understanding

the basic defects and rational

treatment

Structural and functional

characterization of all novel

PROTEINS will unravel new

disease genes.

HOT Areas of Medical Research


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