Summary of protein structure/function

1. The 20 common aa’s have different R groups.2. Amino acids are linked by peptide bonds to form

polypeptides & proteins.3. Proteins are often large & have complex native

(d) structures, but patterns do exist: 1E, 2E, 3E, &4E structures (also motifs, etc.)

4. Native protein structure is maintained by: peptidebonds (1E), weak interactions (2E, 3E, & 4E), anddisulfide bonds ( 3E & 4E).

5. Protein function depends on native structure.1

Amino acids & Protein Structure1. Most jobs (except information storage) in cellsare performed by proteins.

2. Proteins usually have only a few possible stableconformations. How does a protein get from oneconformation to another? Mostly by rotation aboutsingle bonds.

3. Specific protein conformation (structure) isrequired to maintain function.

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N C C

O

O

H

H

H

R

H

aminogroup

carboxylgroup

side chain

I. Functions of Proteins See Biochemistry

II. The α-amino acids (aa) A. Name comes from the structure: The α-C atom

is next to the C'O (carbonyl) C.

1) carboxylic acid group2) α-amino group 3) side chain (a.k.a., R group)4) Circle the α-carbon!

5) Is there a chiral (asymmetric) carbon atom?

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B. Except for glycine (where R = H), the 20common aa all have at least one chiral C atom. Refer to models.

1. L- and D- designations are based on referenceto glyceraldehyde. Biomolecules are linkedthrough biosynthetic pathways. Biochemistsusually go with this designation.

2. The R- & S- designations used in the modelinglab are geometrically based.

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Most α-amino acids have a chiral C atom. Thismeans you can have L- and D- isomers.

L-Alanine D-Alanine5

C. Amino acids (The “20 common”. What doesthat mean?) can be grouped based on similaritiesin the side chains (R group)

1. Non-polar2. Polar but uncharged at normal pH3. Negatively charged (Remember, we are

referring to the side chain.)4. Positively charged

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H2N C C

H

OH

OH

H2N C C

C

OH

OH

H2N C C

C

OH

OH

H H

H

C

C

HH

H

HH

H

H

Gly Ala Val

H2N C C

C

OH

OH

Cys

H2N C C

C

OH

OH

Asp

H2N C C

(CH2)4

OH

OH

Lys

S

H H

H

HH

COOH

NHH

Six of the 20 common amino acids

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D. Why should we care about amino acid sidechains?

1. The side chains play a major role indetermining protein structure/function.

2. Example: Sickle-cell trait is caused by a valinebeing substituted for glutamic acid at only oneposition (out of ~146) in the β-chain ofhemoglobin.

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N C6 C

CH2

O

CH2

C

O-

O

N C6 C

CH

O

CH3

CH3

HH

Glu Val

H H

Importance of amino acid side chain structure inhuman health:

Wild type human Hb has glutamic acid (Glu) atposition 6 of its β-chain. Sickle-cell Hb has Val atposition 6. The remaining 145 amino acid residuesare identical!

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III. Peptide Bond (links monomers to form a polymer)

A. Comment on building big molecules (a.k.a.,macromolecules)

1. Essentially all biological macromolecules arepolymers. Polymers are made by linking alarge number of monomers together: See CD.

n A ö A-A-A-A-A-.....-An-2-An-1-An

2. This is like building a wall out of bricks (asopposed to from stucco).

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C N

C

HC

O C

HCH3

N+

H H

H

H

H

O-

O

peptide bond

Gly-Ala

Atoms shown in redare all in the same plane. Comment re.nature of peptide bond.

3. In proteins, aa are the monomers. They arelinked together by peptide bonds. Note -N-C-C-N-C-C-N-C-C- repeating pattern (seebelow). Which C is (are) the carbonyl-C,which is the α-C?

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4. Nomenclature

a) Two linked aa form a dipeptide (above =glycyl-alanine)

b) Three form a tripeptidec) A moderate number (roughly < 30) form an

oligopeptided) A larger number form a polypeptide

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+H3N CH C

CH

O

CH3

CH3

HN CH C

CH2

O

N

NH

HN CH C

CH2

O

CH CH3

CH3

HN CH C

CH

N

O

OH

CH3

HC C

O

HN CH C

CH2

O

CH2

C

HN CH C

CH2

O

CH2

C

HN CH C

CH2

O-

O

CH2

CH2

CH2

N+

O-O O O-

H H

H

A peptide containing8aminoacid residues (anoctomer)

In 3 letter abbrev: Val-His-Leu-Thr-Pro-Glu-Glu-Lys

Full name: Valyl-Histidyl-Leucyl-Threonyl-Prolyl-Glutamyl-Glutamyl-Lysine

In 1 letter abbrev: V-H-L-T-P-E-E-Y

Can you locate each residue's side chain and determine whether it is non-polar, polar but not charged, acidic, or basic?

Can you make predictions about the solubility in water of each residue's side chain?

Atoms in the peptide backboneare shown in blue.

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5. Peptides can be named by listing their aasequence, starting from the amino terminalend. What is the amino terminal end?

6. The peptide bond exhibits resonance. What isit? (Darth Vader’s voice.)

a) Resonance occurs when there is more than onestable way to arrange the electrons in a moleculeor ion. See next page.

b) Structures with resonance often behave likesomething in between the different resonanceforms.

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C N

H

CaO

Ca

C N+

H

Ca-O

Ca

c) Structures that exhibit resonance tend to be morestable than you would otherwise think.

Peptide bond resonance:1. All six atoms shown are in the same plane. 2. Therefore both the central C and N atoms behave like they

are trigonal planar, sp2 hybridized. (right-hand structure). 3. Note that neither of the structures above provides a perfect

model for all aspects of peptide bond structure/function.

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IV. Primary (1E) Structure of Proteins: thesequence of amino acids

A. Specific proteins in your body have specificsequences. That is, every insulin A-chain startswith Gly the amino terminus, then Ile, etc. (Heterozygosity?)

B. This sequence is called the primary structure ofthe protein. (Polymorphism?)

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V. Secondary (2E) Structure of Proteins A. Secondary structure is regular, repetitive

structure held in place by intrachain HydrogenBonding and comes in two main forms:

1. α-helix

2. β-sheet (two forms of this):

a) parallel [aligned NöC, NöC] b) anti-parallel [aligned NöC, CöN]

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B. Some generalizations:

1. Most proteins have obvious 2E structure.

2. Many proteins have more than 50% of their aainvolved in 2E structure.

C. Specialized 2E structures exist. Example:Collagen triple helix. (proline hydroxylation andscurvy knaves.)

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Part of the A α-helix from humanHb (beta chain)

Can you:1) Trace the peptide backbone?2) Find the amino terminal end? 3) Locate the Hydrogen Bonds that

maintain the α-helical structure?4) Are these Hydrogen Bonds

perfectly aligned?

Would the α-helical structure bemaintained if the Hydrogen Bondswere disrupted?

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C

O

NCHCH

C

R

N

O

CH

H

CN

O

CHC

R

NCH

H

N C

O

OHRHR

H

R

C

O

N CHCH

C

R

N

O

CH

H

CN

O

CHC

R

NCH

H

NC

O

O H R H R

H

R

indicates Hydrogen Bond

Drawing of a β-sheet

Can you:1) Trace the peptide backbones?2) Find the amino terminal ends of the backbones?3) Is this a parallel or anti-parallel structure?4) Are these Hydrogen Bonds perfectly aligned?

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VI. Tertiary (3E) Structure of ProteinsA. Tertiary (3E) describes the location of each of the protein’s atoms in 3-D space. (Re. bends, twists, etc. in secondary structure)

B. Usually, 100% of a given type of protein is in the same 3E structure (Hb, BSE & prions?); this is a very non-random, highly organized situation. If

something is unfavorable in entropy terms (ΔS), there must be a significant amount of bonding (ΔH) holding it in place.

What forces maintain very non-random structures? Remember: ΔG = ΔH ! TΔS ?

1. Peptide bonds (covalent) maintain 1E structure.2. Hydrogen bonds maintain 2E structure.3. 3E structure is maintained by different amounts of a-e different proteins:

a) covalent bonds (!S!S! are a common type)b) hydrogen bonds (??? re. H bonds to solvent)c) ionic bonds (salt bridges)d) hydrophobic interactions (keep the inside on the inside) (actually, mostly a system ΔS term) Comments re. Hydrophobic Collapse & protein foldinge) London Forces Essentially all proteins do b-e in various amounts.

VII. Quaternary (4E) Structure of Proteins(requires multiple subunits)

A. This describes how the subunits fit together.

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B. Examples of proteins with multiple subunits:1. hemoglobin (α2β2)2. insulin (A chain and B chain)3. hCG (β-subunit clinical importance? )

What does hCG stand for? When do (& which?) humans make hCG? hCG structure (pdb: 1hrp) α-subunit: blue; β-subunit: pale green. Red

& bright green atoms are carbohydrate (sugar) molecules that are covalently attached to hCG.

Space-filling model Ribbon model (backbone)

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2E structure is mostly β-sheet with a short α-helix inthe alpha subunit.

VI II. Protein23

Structure re. to Function A. Protein structure has a massive effect on protein

function. Usually alteration in structure radicallyalters (often destroys) protein function.

B. The key to the function of most proteins is thecreation of a unique environment (space) wherecatalysis, transport, or binding can occur.

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IX. Myoglobin & Hemoglobin A. Myoglobin re. O2 storage and diffusion.

1. Myoglobin meaning: myo globin

2. Diving mammals (cetaceans) & myoglobin. See 1mbo protein data bank structure.

Our myoglobin is not used for O2 storage, butto increase the rate of O2 diffusion in ourmuscles.

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B. Hemoglobin (Hb) and O2 & CO2 transport.Maximize O2 binding & release efficiency!!!

1. Hb does have more than 1 stable conformation

a) High O2 affinity form: main form present inlungs (higher pH).

b) Low affinity form: present mostly inextremities (lower pH).

c) Hb shifts back & forth between these formsas it moves through your circulatory system.

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From Principles of Biochemistry, Nelson & Cox, 5th ed. Or go to Google Images,search “hemoglobindissociation curve.”

This graph shows that Hbhas two different forms thathave different affinity for O2.

What percentages of thehigh-affinity form are presentin each of the three curves?

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2. Logic: high affinity form binds O2 in lungs{Hb + 4O2 ö HbC(O2)4}, when HbC(O2)4reaches tissues, there is a shift to the lowaffinity form, and O2 is released {HbC(O2)4 öHb + 4O2}.

3. Other modifiers: bisphophoglycerate (BPG)and CO2 favor formation of low affinity form. That is, they help Hb let go of its O2.

4. Hb also binds CO2 (as HCO3!) and transports it

to the lungs for removal.

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From Principles of Biochemistry, Nelson & Cox, 5th ed.

Left hand curve has ahigher or lower fraction of thehigh-affinity Hb form than theright hand curve?

Which part of yourcirculatory system would be agood match for the centralcurve?

Which part for the righthand curve?

Can you explain why onepart of your circ system wouldbe more acidic?

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From Principles of Biochemistry, Nelson & Cox, 5th ed.

Increasing [BPG] shifts Hbto which form, high-affinity forO2 or low-affinity?

So BPG functions to help Hb .

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Human hemoglobin Backbone: ribbons. One heme associated w/ each subunit: cylinders.

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C. O2 transport and the fetal-maternal unit.

1. One view: Human Hb is really good atbinding O2, but not very good at releasing it.

2. What consequences does this have re. fetus?

3. How do we deal with this problem? a) You didn’t make much Hb β-chain when you were fetal. b) You made a variant of the β-chain called γ.c) Therefore fetal Hb is α2γ2. d) Fetal Hb has higher affinity for O2 than does adult Hb, in

part because fetal Hb does not bind BPG. (So it stays inhigher affinity form.)

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D. Sickle Cell Anemia. (HbS = sickle cellhemoglobin) Thoughts from the group?

1. Small structure change = big function change

2. View structure of wild type (wt) human Hb.

3. Compare sequences (1E structure) of the wt(1a3n) & sickle-cell, HbS, (2hbs) hemoglobinβ-subunit. (Note: yellow-green and sea-foamcolours represent the β-chains in the RasMolrepresentation of 2hbs.pdb.)

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See different pages titled: Comparison of HbA & HbS

Normal (wt) human β-hemoglobin subunit sequence:

1 6 10Val-His-Leu-Thr-Pro-Glu-Glu-Lys-Ser-Ala-.......-His146

Sickle-cell human β-Hb subunit sequence:

1 6 10Val-His-Leu-Thr-Pro-Val-Glu-Lys-Ser-Ala-.......-His146

Only one difference out of 146 aa residues!!!34

4. How does this change alter Hb and rbc function?a) Glu side chain is charged. H2O “likes” to be by it.

b) Val side chain is non-polar. Does H2O like to be by it? Recall that ordered H2O cages are unfavorable (ΔS).

c) During a Sickle Cell crisis (favored by low PO2), separateHbS molecules stick together to minimize Val6 contactwith water. This minimizes cage formation by H2O, but HbS forms insoluble polymers (all kinds of problems).

HbSA(O2)4(aq) W HbS(s) + 4 O2(aq)

What effect would decreasing [O2] have on above equilibrium?(What percentage of Hb is normally in the deoxy form?)

Should sickle-cell patients run wind sprints in the mountains?35

Videos related to blood flow and red cell movement:

The real thing: http://www.youtube.com/watch?v=4yBMY9Wj7z0

Animation: http://www.youtube.com/watch?v=Q530H1WxtOw

Sicle cell crisis: http://www.youtube.com/watch?v=BkC5Hf-AKwo

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5. Why has HbS trait persisted in some populations?

6. How different are human & chimpanzee Hb?

7. How might this relate to human origins???

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X. Dietary Protein & Digestion (Atkins strikes again?)

We can (no longer) make all of the necessary amino acidsfrom scratch (from diet). These aa’s are called

(Should remind you of something?)

A. They are: isoleucineleucinelysinemethioninephenylalaninethreoninetryptophanvaline

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Do you know of a human clinical conditionwhere there is an additional essential amino acid?

Comment on high lysine corn.

B. Example of an amino acid we can make: serine

3-phosphoglycerate ö 3-phosphohydroxypyruvate ö 3-phosphoserine ö serine

Where in our metabolic processes is 3-phosphoglycerate produced?

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