lecture 4' - intro to protein struct i spr08

8/14/2019 Lecture 4' - Intro to Protein Struct I Spr08

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Lecture 4 Introduction toProtein Structure (1)


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Introduction

Proteins are the functional forms ofpolypeptides. represent all levels of the hierarchy of

macromolecular structure (1o

to 4o)

protein structure defined by: chemical properties of the polypeptide chain.

the environment.

Several distinct classes of proteins:

1. globular proteins (water-soluble).2. fibrous proteins (water-insoluble).

3. proteins that associate with membranes.

These differ by tendencies in amino acid sequenceand composition, but can all be described using the same basic principles.


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ro e ns u rom m noAcids

There are 20 common amino acids. all are -amino acids:

amino and carboxylic acid groupsseparated by a single, C carbon.

all are L-amino acids (except Glycine). predominantly zwitterions, at pH 7. distinguished by chemical nature ofR,

the side chain.

Proteins also have other

components: D-amino acids.

e.g., bacterial antibiotics, such asgramicidin.

Covalent modifications, following

synthesis. Disulfide bonds common in eukaryotic


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Adoption of the L-formStructurally Significant

Consider a natural protein, such asRubredoxin: protein in sulfur-metabolizing bacteria.

Fe-S complex shown in green andyellow.

constructed of L-amino acids.

Will Rubredoxin made of D-formamino acids have an inverted

structure? in 1993, L and D-forms of Rubredoxin

were synthesized. structures: X-ray crystallography.

they are exact mirror images.

L- and D- HIV protease also


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Amino Acids

Distinguished by chemical nature of the side-chain. size and shape.

charge.

hydrogen-bonding ability.

ability to form disulfide-bridges, etc.

Amino acids can be broadly classified into 5groups: Aliphatic: R = hydrocarbon side-chain.

Nonpolar: R = other hydrophobic side-chain.

Aromatic: R = aromatic ring.

Polar: R = uncharged, polar group.

Charged: R carries a charge in solution, at pH 7.


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The Hydrophobic Effect

Most proteins are amphipathic: they include both hydrophobic and hydrophilic

residues

Folding generally results in a partitioning of

residues: b/wAq and non-Aq environments. hydrophobic residues

will each desire to avoid water: tend to reside in a membrane or protein interior.

hydrophilic residues will each desire to interact with water: tend to remain hydrated, reside on a protein exterior.

This partitioning b/w Aq and non-Aqenvironments: leads the hydrophobic effect, which drives protein


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The Partition Coefficient

The Partition Coefficient, P: measures the partitioning of a residue betweenAq and

non-Aq environments. Consider a two-solvent system

with separateAq and non-Aq environments;

that are in contact, at equal conditions (e.g.,Temperature).

P answers the question, how much of a given amino acid will reside in each

environment?

For a given amino acid, P is measured by:

P = nonaq /aq, where

i = mole fraction residing in environment i.

conceptually simple, but there are some practical


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Hydrophobicity Scale

Numerous scales of amino acid hydrophobicityhave been proposed. most based on the G

oto transfer from water to

octanol.

which is best is controversial, but one popular scaleis

The Hydrophobicity of Fouchere and Pliska(1983): hydrophobicity parameters, derived from:

transfer from water to octanol. N-acetyl-amino acid amides.

Hydrophobicity parameter for a given amino acid:

= ln P = ln (nonaq / aq)

Then: Hydrophobic: > 0; Hydrophilic: < 0


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Aliphatic Amino Acids

These amino acids have alkyl side-chains: so that R is a hydrocarbon.

All are hydrophobic (P > 1).

hence, have hydrophobicity, = ln P

> 0.Hydrophobicity increases with side-chainlength: = 0.31, 1.22, 1.70, 1.80 for Ala, Val, Leu, and Ile,

respectively.


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Aromatic Amino Acids

All highly hydrophobic. Phe has a hydrophobicity of = 1.79. Tyr is less hydrophobic, at = 0.96,

due to its reactive hydroxyl group.

Trp is the most hydrophobic residue ( = 2.25).Rings bulky and tend to interact with otherrings. due to pi-pi interaction. in Aq. solution, rings

perpindicular. entropically favored.

This is in contrast with

the stacked rings in DNA minimizes solvent

exposure.


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Polar Amino Acids

Each contains groups with partial charges, and therefore, tend to form hydrogen bonds...

Thus, much less averse to water: Asn, Gln, Ser all have negative hydrophobicity values.

= -0.60, -0.22, and 0.04, respectively.

Thr is slightly hydrophobic, at = 0.26. in some scales, Thr is hydrophilic (e.g., in the

hydropathy scale).


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Charged Amino Acids

Amino acids that carry a charge very hydrophilic: Lys and Arg (+) charged at pH 7

very hydrophilic ( = -0.99 and = 1.01, respectively).

His can also be (+) charged at pH 7 (environment-

sensitive). intermediate hydrophobicity ( = 0.13).

Aspartic acid and Glutamic acid (-) charged at pH 7 very hydrophilic ( = -0.77 and 0.64, respectively).


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Overall Charge of a Protein

In passing, we note that the overall proteincharge: depends on the number of acidic and basic

residues...

at the experimental pH of interest.

The Isoelectric Point (pI) = the pH at which the total charge of a protein is

zero.

Protein charge density can be estimated from pI:

c = (pI-pH)/MW

here, MW is the molecular weight of the protein. For pH > pI, overall charge negative (deprotonation).

For pH < pI, overall charge positive.

+ +


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Protein Structure

Proteins may have up to 4 levels of structure: Primary structure:

the N to C amino acid sequence of the polypeptide.

Secondary structure: helices resulting from local folding.

Tertiary structure: global folding of secondary structures into a larger

structure.

Quaternary structure: association of several, independent polypeptides.

We will look at each, in detail 1o and 2oStructure (this Lecture)

3o

and 4o

structure (next Lecture)


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Protein Primary Structure

A polypeptide - covalently linked chain of aminoacids. each linked amino acid called a residue. each pair of residues connected by a peptide bond:

The N-terminal to C-terminal sequence ofresidues: Is the primary structure (1

o) of the encoded protein.


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Anfinsens Principle

Anfinsens Principle is basic to biochemistry: The information needed to fold a macromolecule into

its native, 3-D structure is contained in its sequence. Denatured ribonuclease spontaneously refolds into the

enzymatically active form, in vitro (Anfinsen, 1963).

1o

structure then specifies the higher structure: Each sequence corresponds to a well-defined 3-D

structure. or to a family of closely related structures with activity.

the native state.

On the other hand a unique structure does notrequire a unique sequence. level of sequence homology required for similar

structure is only about 25%-30%.

non-homologous sequences can also have similar


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Traditional Names for Helices

Determined by the nature of the repeating H-bond: N = number of residues in 1 helix turn.

d = number of atoms in the ring formed by each H-

bond donor (amino-H) and acceptor (keto-O).

each helix assigned the name, Nd.

e.g., in a 310 helix,

Each turn contains N=3 residues;

each H-bonded donor/acceptor pair form a ring of d =10 atoms.

This notation has several shortcomings: a -sheet cannot be described in these terms:

each strand is a 2-fold helix, butH-bonds between

strands.


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The Standard Terminology

For displacements down the helical axis: total rise/turn = pitch, P.

rise/step = helical rise, h.

steps/turn = helical repeat, c (= n).

P = c h

For angular displacements about the axis: angle/step = = helical angle, or twist.

= 2/c = 2 h/P

For helical symmetry about the z-axis, Positions of adjacent steps then related by:


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Helix Handedness

The Symmetry Matrix does not uniquely specify ahelix: a helix can be left or right-handed.

in principle, handedness given by : right-handed: > 0.

left-handed: < 0.

but we have the convention: >= 0. all rotations defined as right-handed.

Our notation is degenerateWe distinguish right and left helices by: the axial displacement: P or h.

For right-handed helices, P and h > 0.

For left handed helices, P and h < 0.

The helix shown here is right-handed.


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Naming Helices by Symmetry

Helical symmetry is denoted by NT:

N denotes its N-fold rotation operator. T = translation generated by the symmetry

operator, in repeats (monomers).

compare with Nd notation.

Example: Take the helix at right... Each residue rotated by +120

o.

3-fold helical symmetry (N = 3).

1 application of the symmetry operator: translates a residue by +1 repeat, or P/3.

thus, T = 1.

This helix has 31

symmetry.

note that it is a right-handed helix.

ri ht-handed helices with inte er N are N helices.

e e x as


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e 10 e x as 1Symmetry

Example: The 310 Helix

one of the two common helices inproteins.

Commonly named by Nd notation:

N=3 residues/turn.

d=10 atoms in each H-bond closedring.

note: green line includes a sharedH.

ith keto-O H-bonded with (i+3)thNitrogen.

The 310 helix has 31 helical

symmetry.

Each residue rotated by +120o.

T e e x as 18


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T e - e x as 185

Symmetry

The most common helix in globularproteins. i

thketo-O H-bonded with the (i+4)

thNitrogen.

13 atoms b/w H-bond donor and acceptor.

The -helix has 3.61 helical symmetry.

Each residue rotated by +100o.

3.6 residues/turn (N = 3.6).

1 application of the symmetry operator:

translates a residue by +1 repeat, or P/3.6. thus, a right-handed helix, with = 1.

For helices with non-integral symmetry: N and T converted to integers

-helix said to have 185 symmetry. 18 residues in 5 full turns.


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The trans-conformation is a 21

helix

Consider the fully extendedpolypeptide: keto-O and amino-H of each Peptide bond in

the trans configuration, and

For each residue, both and = 180o.

remember our convention: Polymer chemistry: cis = 0

o.

Biopolymer in a fully trans conformation

Somewhat expanded use of the term, trans. since cis/trans defined for confi urations.


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The -strand is also a 21 Helix

Differs from the trans-conformation: a -strand is pleated, like a curtain.

shorter (smaller rise, h).

In order to be stable, -strandscombine to form sheets: by strand-to-strand hydrogen

bonding.

two general types: anti-parallel sheets (A).

parallel sheets (P).

Generally, these sheets are

twisted.

Standard Helices of


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Standard Helices ofBiopolymers

these include all of the standard, 2ostructures of

biopolymers.

Thus, our original contention is correct: the only symmetric building block formed by a chain

of chiral units is a helix.

Question: can the backbone adopt thesestructures?


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The Peptide Bond

The peptide bond links each pair of adjacent residues; is an amide linkage,

with a partial double-bond character.

this bond not freely rotating: 6 atoms constrained to a plane:

Ci, Ci, Oi, Ni+1,Hi+1,Ci+1.

The peptide bond can adopt one of 2configurations: based on the positions of Ci and Ci+1. The trans-configuration (i= 180

o):

energetically favoredusually adopted.

The cis-configuration: (i= 0o):

sterically hinderedenergetically unfavorable. exception: Proline (cis and trans nearly


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The Ramachandran Plot

The Ni-Ci bond, and Ci-Ci bond are single bonds: in principle, each may rotate freely

and could then assume any values b/w +/- 180o.

however, this is true only for Glycine (R = H).

For other R-groups, non-covalent interactions b/wadjacent side-chains: place energetic constraints on and .

thus, some conformations (,) are sterically disallowed.

Sasisekharan and Ramachandran (1968): first plotted the van der Waals energies of interaction vs.

(,). using poly-L-Alanine

R = Me, the minimally constrained group (with a C-Carbon).

with the trans-configuration for each peptide bond.

the resulting plot is a Ramachandran Plot.


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The Ramachandran Plot(cont.)

Plot shown in terms of allowed regions here, c = helical repeat, with +/- = right/left-handed.

dark regions = sterically allowed. lightly shaded regions = moderately disfavored.

others = disallowed.Torsion angles of helices:

all in allowed regions. Note: L- left handed -helix.

Glycine only (achiral).

Not a good model for: Glycine or Proline. residues w/ bulky side-chains. more on this in Lecture 8.

esp. sequence effects.


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Conclusion

In this Lecture, we have discussed: Amino Acid Residues

Characteristics and Hydrophobicities

Protein 2o

structure:

The local, helical structures adopted by polypeptides.

We will continue our discussion in Lecture 4: The intermediate level structure of Proteins:

Super-2o

and Domainal structure of Proteins. Methods of Visualization of the overall 3-D structures

of Proteins. Protein 3

ostructure:

contact plots.

Protein 4o

structure.

lecture 4' - intro to protein struct i spr08

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