II. Patterns and forms in protein structure

2. Patterns and forms in protein structure

•Helices and sheets

•The hierarchical nature of protein architecture

•Structure based classification of proteins

•protein folding: Intra-cellular pathogens and the survival of the flattest

•Protein folding and disease: Amyloidoes, Parkinson, Huntington, Prion disease

Protein Secondary Structure

-helix -sheet

These secondary structures are highly present in proteins due to:

-They keep the main strain in an unstrained conformation

- Satisfy the hydrogen-bonding potential of the main-chain N-H and C=O groups

These secondary structures link in a specific way in different combinations to perform the final protein structure

-helices are formed from a single consecutive set of residues in the amino acid sequence

The H-bond links the C=O group of residue i with the H-N group of residue i + 4

There are alternatives to the helix configuration giving more constrained or less constrained structures:

-310 helices, in which hydrogen bonds form between residues i and i + 3

- -helices, in which hydrogen bonds form between residues i and i + 5

This configurations are much rarer due to the constraints and effects they have on the protein stability.

-sheets are formed by lateral interactions of several independent sets of residues.

They can bring together sections of the chain widely separated in the amino acid sequence

In this figures, all the strands are anti-parallel

Tertiary and quaternary structure

Tertiary structures are the result of the different combinations of helices and sheets

The different combinations lead to different spatial arrangements and different patterns of interactions between amino acids of helices and sheets. This will be the basis for the so called FOLDING PATTERN

Many proteins contain more than one subunit, or monomer. They may be multiple copies of the same polypeptide chain, or combinations of different polypeptide chains which assembly form the QUATERNARY STRUCTURE

Protein stability and denaturation

The native structure of proteins can be broken up, by heating or by high concentrations of certain chemicals such as urea (DENATURATION)

Denaturation destroys the secondary, tertiary and quaternary structures but leaves the polypeptide chain intact.

The stability of the the main chain will ensure that, ones natural conditions restored, the protein will acquire the normal productive folding conformation and thus its function.

Proteins are only stable under very narrow conditions of solvent and temperatures. Breaking these conditions will break the intimate intramolecuar interactions, will change the main configurations of the backbone and will lead to non-productive conformations

Giving the changeability of these conditions, the cell has developed many mechanisms to buffer these effects (Moran et al. 1996; Fares et al. 2002 a, Fares et al. 2002 b, Fares et al. 2004).

Productive protein conformation

The protein conformation ensures the intra-molecular interactions that are essential for forming the active sites and therefore for enabling the protein to have a biological activity.

Active sites in enzymes only require 10% of the total number of amino acids in the protein. The different molecular interactions between different local secondary protein structures have the role of:

- Scafolding to enable the appropriate conformation for the formation of the active site

- enable conformational changes as part of the mechanism activity (Steroid Hormone receptors)

- Some residues are in strained conformation playing an important role in catalysis

Due to the crowded cell environment, slow-folding proteins tend to aggregate non-specifically leading to several known diseases:

Alzheimer, Prion disease

The role of chaperones is essential in ensuring correct protein folding

Protein structure and conformation

5. Proteins are polymers containing a backbone or a main chain of repeating units (peptides) with the main chain attached to it

-N-C-C-N-C-C-........

O O

Si-1 Si

Amino Acids

Asymmetric carbon

Amino acids vary in

size

Hydrogen-bonding potential

charge

Amino acids are chemical building blocks

HK

R

E

D

F YW

IL

V

M

A

Q

S N

T

CH

G PC

S-S

polar

positive

negative

charged

beta-branched

aliphatic

aromatic

hydrophobic

What amino acids look like

C

R

HN C

O

OH

H

H

side chain

aminogroup

carboxyl group

Sidechain nomenclature

C

X

X

X

carbon alpha, central chiralcarbon of the amino acid

beta;first sidechain position

gamma position

delta position

epsilon position

zeta position X

X

Xeta position

Small amino acids

Gly: G

Ala:A

Cys:CSer:S

Asp: D

Pro: P

Aliphatic amino acids

Ala:A

Ile: I

Leu: L

Val: V

-branched amino acids

Ile: I

Thr: T

Val: V

Aromatic amino acids

Tyr: Y

Trp: W

Phe: F

His: H

Polar amino acids

Asn: N

Gln: Q

Ser:S

Thr: T

Tyr: Y

Positively charged amino acids

Arg: R

Lys: K

His: H

Negatively charged amino acids

Glu: E

Asp: D

Chirality

• Amino acids are not flat and two dimensional!

• Groups are arranged around the central carbon atom in a tetrahedral fashion (why?)

• There are two possible ways for the groups to be arranged:

Amino acid chirality

C

R

HN CO

C

R

HNCO

L-form D-form

amino acids inproteins are almostalways in the L-form

D-form occurs rarely --peptide antibiotics, somepeptide toxins

Peptide chemistry

C

R

HN+ C

O

O-

H

HC

R

H

N+ C

O

O-H H

H

amino acids dissociate in aqueous solutionto form a zwitterion (ionic species with twoindependent charged groups)

Peptide chemistry

C

R

HN C

O

H20

H

H C

R

H

N C

O

OH

H

peptide bond

the amino acid polymer formswhen the carboxyl group of one amino acid condenses withthe amino group of the next

Protein folding

The energy of protein conformation depends on:

Interaction of sidechains and main chains

Interaction with solvents and ligands

The environmental conditions of the cell

Native state

Proteins follow the shortest temporal and energetical pathway to acquire the most stable conformation

DenaturedSpontaneous aggregation

Non-specific aggregates

Chaperones

Functional conformations

Protein Folds: sequential, spatial and topological arrangement of secondary structures

The Globin foldThe Globin fold

DnaK DnaJ

mRNA

protein

a

bc

A

GroEL

GroEL

ATP and GroES binding

ADP + Pi and

GroES release

BGroES

t = 1

t = 2

t = 3

t = 13512 lines REL4548 (MAF)12 lines REL7550 (MXR)

Vertical transmission of E. coli as a simulating system of endosymbiosis

Comptenece experiments

Day –1: adapt to DM25 ( 3-5 replicaqtes)

Day 0: mix both competitors 1:1, determine their proportions

Day +1: determine proportionsof competitors, estimate W

Day –2: grow on LB every competitor

ara ara+

BamHI

PP

HindIII HindIII

Pbla

tetA tetRyjeH

groE

SalI XhoI

S L

0.4

0.6

0.8

1.0normalmutator

Ancestral evolved groEc

W

groES groEL

137 Å

57 KDa146 Å

Apical domain

Y199, S201, Y203, F204, L234, 237, 259

V263, 264

Intermediate domain

Equatorial domain

GroEL as a compensatory mechanism

95100

100

100

99

100100

96

99

97

95

0,05

A. pisum PS

S. Avenae PS

M. Persicae PSR. padi PS

S. graminum PS

P. populeum PS

T. caerulescens PS

C. leucomelas PS

T. salignus PS

T. suberi PSE. carotovora

E. coli

S. typhimurium

E. aerogenes

0.05

100

9999

93

100

B. germanica PS

E. libidus PS

P. americana PS

B. orientalis PS

L. dicipiens PS

B. gingivalis

P. gingivalis

E. coli

R.maidis PS

R.padi PS

S.graminum PS

M. persicae PS

S. avenae PS

P.populeum PS

C. Leucomelas PS

T. caerulescens PS

B. pistaciae PS

T. suberi PS

T. salignus PS

W. glossinidia PS

B. tabaci PS

A. proteus PS

E. carotovora

K. pneumoniae

E. aerogenes

S. enterica

S. typhimurium

S. glossinidia SS

S. oryzae PS

P. putida

P. aeruginosa

P. gingivalis B. gingivalis

L. dicipiens PS

100

99

100

100

100

100

10061

9910064

97

99

100

100

100

100

94

100

99

94

100

77100

42

0.1

A

B

Flavobacteria E. libidus PS

B. germanica PS B. orientalis PS

P. americana PS100

93

CD

E

FH

GI

J

-proteobacteria

Branch Average

A 21.98

B 13.45

C 1.57

D 1.37

E 2.58

F 1.42

G 3.93

H 3.98

I 3.68

J 4.035

Positive selection in the endosymbiont GroEL

A B

Convergent adaptive evolution in GroEL from endosymbiotic bacteria

Protein structure stability and its ability and specificity to bind ligands depend on different chemical forces:

Hydrogen bonding

Covalent bonds

Hydrophobic effect

Conformation of polypeptide chain

Condensation of amino acids produces a polypeptide chain, with the backbone atoms linked through the peptide bond

The angles of internal rotation around the bonds determine the pattern of protein folding

Simple bonds not restricted by the electronic structure but by esteric collisions

The double bond character of the peptide restricts internal rotation

The peptide group occurs in cis and trans forms, being trans more stable for all amino acids except for Proline

All the cis forms in a polypeptide are restricted to Proline and the amino acid preceding it due to the small difference in energy between cis and trans

The dominance of the trans peptide bonds determines two angles for the main chain conformation of each residue and , being some of their combinations disallowed from the energetic point of view

= -125º, and = +125º

The Sasisekaran-Ramakrishnan-Ramachandran diagram