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A new approach towards deciphering the protein code: The protein assembly model. Claire Lesieur [email protected] Membrane (Lipids). Proteins. Nucleus (chromosome). Elements of the living world. Protein. Nucleus. Lipids. DNA. CHON. Chromosome. Protein Biological activities.

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A new approach towards deciphering the protein code the protein assembly model

A new approach towards deciphering the protein code: The protein assembly model

Claire Lesieur

[email protected]


Elements of the living world

Membrane (Lipids) protein assembly model

Proteins

Nucleus

(chromosome)

Elements of the living world

Protein

Nucleus

Lipids

DNA

CHON

Chromosome


Protein biological activities
Protein Biological activities protein assembly model

  • Cutting

  • Recognition

  • Enzyme

  • Signaling

  • Carrier

  • Shape generator

  • Road networks


Structure function relationship in proteins
Structure-function relationship protein assembly modelin proteins

  • Function

  • Shape

  • How the shape provides a particular function

  • How the shape is acquired


? protein assembly model

?

GKKHDGATTYQW


The protein folding problem
The protein folding problem protein assembly model

  • How it folds: Mechanisms of protein folding

  • How the information is encrypted in the sequences: CODING problem

ADRTGGILLKMHGGARECVVP


All the information necessary for the protein folding is within the protein primary sequence

C.B. Anfinsen, Haber, E., Sela, M. & White, F. H. , Proc. Nati. Acad. Sci. USA 47 (1961) 1309-1314.

Levinthal’s paradox(1968): not random search but directed

Levinthal, C. (1968) J. Chim. Phys. 65, 44-45.


Mechanism

COOH within the protein primary sequence

H2N

s-hours

ms

Structure Tertiaire

Structure primaire

Structure Secondaire

Mechanism

Short range interaction

long-range

interactions

short-range

interactions


Code still unknown
Code: still unknown within the protein primary sequence

X-ray crystallography + NMR: PDB

3D modeling: PDB

~ 70 % Sequence similarity: 3D modeling

70 % similarity: different shape

Low sequence similarity: similar shape

Amino acids on the surface of proteins: changeable


Transmembrane domains of membrane proteins
Transmembrane within the protein primary sequence domains of Membrane proteins

b-strands transmembrane domain: 1010101

a-helicetransmembrane domain: 11111111111111111


Biologically active amino acids
Biologically active amino acids within the protein primary sequence


Sequence shape predictions
Sequence-Shape predictions within the protein primary sequence

  • Geometrical constrain

  • Chemical constrain


To read sequences you need to determined comparable sequences
To read sequences you need to determined comparable sequences

  • Domains

  • Shape and role

?

Sequence Pattern

?

Sequence Pattern


Protein assembly
Protein assembly sequences


Aerolysine
Aerolysine sequences

Trends in Microbiology (2000). Vol 8 (4):169-172


ER sequences

Cholera toxin

CtxA

CtxB5

  • AB5 toxin

    • A catalytic subunit

    • B receptor binding subunit

  • GM1: cell receptor

  • Endocytosed and traffic to the ER

  • ADP ribosylation of Ga subunit

  • Increase of cAMP leading to water loss



Assembly sequencesin vitro

pH 7

pH 1

15 min

Pentamere

Monomere


2d structural level short range interaction
2D structural level: short range interaction sequences

5

2 10

0

5

-2 10

5

-4 10

pH 1

5

-6 10

Mean residue Molecular Ellipticity

pH 7

5

-8 10

Native

6

-1 10

6

-1,2 10

200

210

220

230

240

250

Wavelength (nm)


3d structural level long range interaction
3D structural level: long range interaction sequences

  • Trp-fluorescence

300

lex= 295 nm

lem=352 nm

Fluorescence Intensity (a.u.)

200

Fluorescence Intensity

100

unfolded

0

Time (min)

320

340

360

380

Wavelength (nm)


Functional test
Functional test sequences

His

CtxB

100

80

Function

60

HISTIDINE

40

20

4,5

5

5,5

6

6,5

7

7,5

8

0

pH


sequences

CtxB5


LTB sequences

CtxB

Cholera toxin B

Heat labile enterotoxin B


N-terminal sequences

100

LTB

CtxB

80

Function

60

N-terminal

40

20

0

4,5

5

5,5

6

6,5

7

7,5

8

pH


LTB5 sequences


Kinetics differences sequences

On pathway intermediates differences

It is particular amino acids that are responsible for each individual step of assembly and folding


Fundamental question
Fundamental question sequences

  • Alzheimer, Parkinson, Prion diseases

Protein X: FOLD state: healthy

Information for interfaces

(Protein X)n: Assembly state: Lethal


Theoritical approach
Theoritical sequences approach

  • Protein Interface formation

  • Rules?

  • Mechanism?

  • Preferential geometries related to preferential sequences of amino acids?


INTERFACES: sequences

Zone de contact entre monomeres voisins


Analyses des interfaces
Analyses des interfaces sequences

Interface Trimer pentamer heptamer

Brin 1

Brin 2

0101 0101 Ch111Ch

n.a. Ch111Ch 1111/1


Trimeric domain
Trimeric sequences Domain



Oligomeric proteins
Oligomeric sequences proteins

Nombre de monomer 2 3 4 5 6 7 8 9 10 11 12

Nombre de cas 5722 1035 2340 168 721 46 512 45 87 8 205


Programme detection protein interfaces
Programme sequences detection Protein Interfaces

Monomer M

513 -524

LMITTECMVTDL

aaa-bbbbbbb-

Monomer M+ 1

35-49

GRNVVLDKSFGAPTI

--bbbb-------bb

Distances


2HY6 (30) sequences

1 30

beta

1N9R (68)

19 86

alpha

1WNR (94)

1 94

a+b

2F86 (129)

344 471

1JBM (78)

10 88

rc

1G31 (107)

5 111

1LNX (74)

8 80

1Q57 (483)

64 549

2RAQ (94)

3 97

1GRL (518)

6 523

1IOK (524)

2 526

1PZN (240)

96 336

1J2P (229)

4 233

1Y7O(194)

1 194

2F6I (189)

177 367

1TG6 (193)

1 193

2CBY (179)

15 194

1OEL (525)

2 525

1LEP (92)

1 92

3BDU (51)

2 53

1HX5 (92)

5 97


Putative lipoprotein from e carotovora
PUTATIVE LIPOPROTEIN sequencesfrom E. CAROTOVORA

3BDU 20-29, 38-53


Common protein interfaces of unrelated proteins
Common protein interfaces of unrelated proteins sequences

3BDU 1--111011-110110--10

1G31 0--1-1001-100100--00

1JBM 11001000101100101101

1LNX 1--0100010110000---1

1N9R 0--0100011110010--11

1J2P ----1000101100101--1

1HX5 ------0011110010--11

1LEP 0---10001000--00--11

Con2 ----1-001-1100-0-


1LEP: sequences1-8, 88-94, 40-57

1WNR: 1-8, 88-94, 44-57, 62-77

1HX5: 5-11, 94-97, 51-62, 68-80,27-30

1G31: 8-15, 104-111, 68-85


1N9R sequences

yeast

Methanobacterium Thermautriophicum: extremophile

1JBM

P. aerophilum: bacterium

1LNX


1 sequences

yeast

1 + 1

Methanobacterium Thermautriophicum: extremophile

1JBM: 12-18, 42-50, 64-83

1 +1 +1

1N9R: 66-82

P. Aerophilum

Hyperthermophilic bacterium

1LNX: 10-15, 25-32, 40-48, 63-77


2CBY sequences


Conclusion
Conclusion sequences

  • Geometry and function related

  • Family of protein interfaces

  • Assembly keys


Future
Future sequences

  • Classification of protein interfaces: Database

  • Systematic analysis of protein interfaces-subjective classification

    • Mathematical approach: Laurent Vuillon (LAMA)

  • Functional analysis of protein interfaces

    • Protein Assembly mechanism from block: Giovanni Feverati

    • Stoechiometry/Symmetry: Paul Sorba

    • Experimental tests: Claire Lesieur


  • Acknowledgment
    Acknowledgment sequences

    • Alicia Ng Ling

    • Mun Keat Chong

    • Boon Leng Chua

    • Danyang Kong

    • Giovanni Feverati

    • Paul Sorba


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