Lecture 7 & 8:PRO

TEIN A

RCHITECTU

RE IV:Tertiary and Q

uaternary Structure

Margaret D

aughertyFall 2004

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right-handed α-helix

left-handed α-helix

Point your thumb up in the

direction of the α-helix(N

-->C).

Look where the “base” of thehelix spirals are. If they

match the knuckles on yourright hand it is a right-

handed helix.

----------or-----------

If the helix spirals up in acounterclockwise direction, it

is a right-handed helix.

How to tell a left- vs. right-handed a-helix

Tertiary Structure

•Tertiary structuredescribes how the

secondary structureunits associatewithin a single

polypeptide chain togive a three-dim

ensionalstructure

flavodoxin

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formation of large num

ber of intramolecular hydrogen bonds

reduction in hydrophobic surface area from solvent

Tertiary Structure: Basic Tenets - the “truths”1). A

ll information for folding is contained in the prim

ary sequence.2). Secondary structure form

ation is spontaneous - a consequence of theform

ation of hydrogen bonds.3). N

o protein is stable as a single layer - hence secondary structuralelem

ents pack together in sheets.4). Connections between structural elem

ents are short - minim

ization ofdegrees of freedom

- keeps structures compact.

Consequences1). Secondary structures are arranged in a few com

mon patterns - i.e,

resulting in protein “families”.

2). Proteins fold to form the m

ost stable structure. Stability arises from:

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All alpha

(human growth horm

one)

Note that som

eparts of a proteinstructure are not

regular (i.e., helical-like or sheet-like) .

These are oftenreferred to asdisordered or

random coil regions.

However a betternom

enclature is“natively random

”.A

ll beta(retinol binding protein)

Tertiary Structures

Alpha-beta barrel(triose isom

erase)

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Tertiary Structures

Globularproteins:

compact structures;

different folds fordifferent functions

Mem

braneProteins:

found associatedwith various

mem

brane systems

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Fibrousproteins:

Filamentous; playa m

ajorstructural role in

cells & tissues

Fibrous Proteins•

Share properties that give strength &/or flexibilityto the structures in which they occur;

• Fundam

ental unit is a simple repeating elem

ent ofsecondary structure;

• Insoluble in water; large percentage of hydrophobicam

ino acids;•

Usually the hydrophobic surfaces are hidden in the

elaborate supramolecular com

plexes;•

mechanically strong ; perform

important structural

functions•

Strength is enhanced by cross-links (disulfidebonds).

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Secondary Structures andProperties of Fibrous Proteins

Structu

reC

haracteristicsExam

ples ofoccu

rrence

α-H

elix, Cross-linked

by disulfide bondsT

ough, insolubleprotective structures ofvarying hardness andflexibility

α – K

eratin of hair,feathers and nails

β-Conform

ationSoft, flexible filam

entsSilk fibroin

Collagen triple helix

High tensile strength,

without stretch

Collagen of tendons,

bone matrix

FIBROUS PRO

TEINS: α-Keratin

What: Part of the “interm

ediate filament proteins” which have m

ajorstructural roles in nuclei, cytoplasm

and cell surfacesW

here: Found in hair, fingernails, claws, horns, animal skin

Composition: Long stretches of α-helices (> 300 residues)

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Coiled-Coils

•Interactions are stabilized byhydrophobic interactions between the α-helices;

•H

eptad repeat (a-b-c-d-e-f-g)n where a &d are nonpolar & lie in the center of thecoiled coil;

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Evolved for strength; helicalnature confers flexibility

Coiled-coil is a “super twist”left-handed helix

Distortion of helix to 3.5

residues/turn

Hydrophobic faces interacting

in a close interlocking pattern

α-keratin:

Contact side chains (red balls) interlock

FIBROUS PRO

TEINS: β-Keratin

What: Part of the “fibroin proteins”

Where: silk, bird feathers

Composition: stacked anti-parallel β-sheets; strength

Sequence: Alternating G

ly-Ala/S

er

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Ala/Ser faces

interact withone another

Gly faceinteracts withanother glyface

COLLA

GEN

What: Greek for glue; defined as “that

constituent of connective tissue whichyields gelatin on boiling”W

here: Principal component of

mam

malian tissue; constitutes ~25%

ofa m

amm

als protein content; more than

30 varietiesCom

position: Triple helixSequence: Gly-X

-Y; X usually Pro, Y

usually Pro/HyPro

> 3000Å long; 15Å

in diameter

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Collagen Primary Structure

•A

pprox 1000 AA

/chain•

Repeats of Gly-X-Y where X

is often Pro and Y isoften hydroxyproline or proline

Composition

G ~ 35% A

~ 11% P/H

P ~ 30%

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MO

DIFIED

AM

INO

ACID

Spost-translational m

odifications add functionality to amino acid

Hyp: stabilizes

tropocollagen viaintrachain H

-bonds

Hyl: stabilizes fibrils

via its ability to cross-link; attachm

ent ofCH

O groups

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Consequences of Collagen Primary Structure

Distortion of backbone

due to high content ofglycines and prolines

Can’t form “norm

al”secondary structures

Forms triple helix

Every third residuefaces inside

Interior is compact;

hence interior residueis glycine

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Consequences of Collagen Primary Structure

Fit occurs because Glystrand1 lies adjacent to X

strand2 and Ystrand3

Stabilization from hydrogen bonds

Glystrand1 N

-H to X

strand2 C=O hydrogen bond

Hydroxyproline form

s hydrogen bonds BIO

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•Folding of 2˚ structural elem

ents;•

Side chain

location varies

withpolarity;–

Non-polar are inside

• (A

, V, L, I, M & F);

–Charged on the surface•

(D, E, K, R, H

);

–U

ncharged polar mostly surface,

but interior as well•

(S, T, N, Q

, Y & W);

–N

early all H-bond donors have an

H-bond acceptor;

–Interior

of a

structure is

TIGHTLY packed;

•3˚ structures are frequently built ofdom

ains.

GLOBU

LAR PRO

TEIN

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Globular Proteins: 4 major classes

I: antiparallel α-helixPacks in bundles; Left-handed twistU

sually regular, uniform; U

sually 4-helix bundles

Globin proteinsII: Parallel or m

ixed β-sheet proteins parallel β-sheetshave hydrophobes on both sides of sheet ---> thesem

ust be core structuresIII. A

ntiparallel β-sheet structures antiparallel β-sheetshave hydrophobes on one side of sheet and polarresidues on the other side. These β-sheet structurescan be surface exposed.

IV: Metal & D

isulfide rich proteins Small, < 100 a.a.

Structure dependent upon either the metal or disulfide

CORES O

F PROTEIN

S: α-HELICES A

ND β-SH

EETS

Ribonuclease ACitrate synthase

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GLOBU

LAR PRO

TEINS A

ND α-H

ELICES:return of the helical wheel!

SURFA

CE HELIX

: AM

PHIPA

THIC

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GLOBU

LAR PRO

TEINS A

ND α-H

ELICES:return of the helical wheel!

INTERIO

R HELIX

: HYD

ROPH

OBIC

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GLOBU

LAR PRO

TEINS A

ND α-H

ELICES:return of the helical wheel!

SOLVEN

T-EXPO

SED H

ELIX: PO

LAR/CH

ARGED

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β-sheets: an example of IFA

BP

Hydrophobic = green

Charged = red

Looking in cavityfrom

bottomleft of the 3front sheets

pdb file: 1AEL

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POLYPEPTID

E CHAIN

S HAVE A

RIGHT-H

AN

DED

TWIST

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SINGLE “LA

YER” STRUCTU

RES ARE N

OT STA

BLEBIOC 205

1). Proteins aretightly packed

GLOBU

LAR PRO

TEIN STRU

CTURE: other considerations

2). Proteins havenatively randomstructure

3). Proteins haveflexible segm

entsBIO

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Proteins Are D

ynamic:

X-ray vs N

MR Structures

A chain of insulin

Solution structure“fam

ily” of structuresStatic structure

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MOTIO

NS CO

VER A RA

NGE O

F TIME: seconds to fem

toseconds

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MULTIM

ERIC PROTEIN

S

•Q

uaternary structuredescribes how two or

more polypeptide chainsassociate to form

anative protein structure

(but some proteins

consist of a singlechain).

Hem

oglobin

Hb tetram

er: 2 “alpha chains” and 2 “beta chains”BIO

C 205BIO

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CLOSED

QU

ATERN

ARY STRU

CTURES

The association stiochiometry is finite

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Isologous vs. Heterologous A

ssociations

How the packing interfaces of the m

onomers com

e together

OPEN

QUATERN

ARY STRU

CTUREBIO

C 205

tubulinH

IV-1 particle

Tobacco mosaic virus

OPEN

QUATERN

ARY STRU

CTURE

Coat protein40 kD

RNA

genome in red

surrounded by helical array of subun its300 nm

x 18 nm2134 copies of coat protein!

STABILIZA

TION O

F QUATERN

ARY STRU

CTURE

Quaternary Structure are

Stable!

Kd = 10

-8 to 10-16 M

or50 to 100 kJ/m

ol instability!

alcohol dehydrogenase

Chemical interactions offset loss of entropy

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WHY H

AVE Q

UATERN

ARY STRU

CTURE:

ADVA

NTA

GES OF H

AVIN

G ASSO

CIATIN

G SUBU

NITS

Subunit association can shieldhydrophobic groups from

water

Reduction in surface to volume

ratio: we will see that hydrophobicinteractions in the protein core have agreater im

pact on protein stability thansurface hydrogen bonding with water

Bringing catalytic sites together: important for proteins with m

ultiplecatalytic functions - m

ore efficient in terms of “handing off” substrates from

one site to another for subsequent chemistry.

Genetic Economy and

Efficiency: less DN

A is used to

code for a protein

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Cooperativity: Binding of asubstrate to one subunit caninfluence the binding of asecond substrate to anothersubunit.

QUATERN

ARY

STRUCTU

RESCA

N BE

SIMPLE O

RCO

MPLEX

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REVIEW1). Tertiary structure describes the three-dim

ensional structure of a polypeptidechain.2). The 3 m

ajor classes of 3o structure are fibrous proteins, globular proteins, and

mem

brane proteins.3). Fibrous proteins are hydrophobic proteins that give strength and flexibility.4). Coiled-coils are stabilized by hydrophobic interactions.5). Globular proteins constitute the m

ajority of proteins , consist of α-helices andβ-sheet and have a hydrophobic core.6). Polypeptide chains have a right-handed twist.7). Globular proteins have layers.8). Globular proteins are densely packed9). Globular proteins can have flexible regions.10). Proteins display m

otions (returning to the idea that life is dynamic!)

11). Quaternary structures describe the association between polypeptide chains.

12). Quaternary associations can be “open” or “closed”

13). Quaternary structures are stable (an interplay between entropy and chem

icalinteractions) and confer certain advantages to an organism

.

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