Protein Sectors: Evolutionary Units of Three-Dimensional StructureNajeeb Halabi, Olivier Rivoire, Stanislas Leibler, and Rama Ranganthan

Cell 138, 774-786, August 21, 2009

Journal Club

Yizhou Yin

Sep 23, 2009

Sequence Conservation“…sequence conservation – the degree to which the frequency of amino acids at a given position deviates from random expectation in a well sampled multiple sequence alignment of the protein family...”

sequence structure property/functionevolution

sequence conservation

Evolutionary relationship

Structural/functional importance

Hypothesis

-However, in the 3-dimensional structure of protein, the large amount of interactions between amino acid residues are also fundamental “structural elements”.

-Amino acid distributions at individual position should not be taken as independent of one another.

-Investigation of correlations between sequence positions in protein family leads to decomposition of the protein into groups of coevolving amino acids – “sectors”.

Hypothesis: the sectors are features of proteins structures and reflect the evolutionary histories of their conserved biological properties.

S1A Family

Serine protease

Clan

SASB…

Family

S1S2…

Sub-family

S1A trypsinchymotrypsintryptasekallikreingranzyme

…

Broad distribution and functions

Prokaryotes

Invertebrates

Vertebrates

Digestion

Blood clotting

Inflammation

…Binding site - specificity

Catalytic triad – active site

Member

…rat trypsin (3TGI)

Method Outline Identification of sectors

Statistical Coupling Analysis

Statistical Independence Correlated entropy

Physical connectivity

Distinct biochemical properties Alanine mutagenesis Catalytic power & thermal stability assays

Independent divergence Sequence similarity analysis

From Sequence to SectorsMultiple sequence alignment of 1470 members of the S1A family (single domain)

NCBI nonredundant database through iterative PSI-BLAST

Alignment: Cn3D, ClustalX

Standard manual adjustment methods

Di(a): Divergence (or relative entropy)

fi(a): Observed frequency of amino acid a at position i

q(a): Background frequency of a in all proteins

Position Conservation

SCA matrix (conservation-weighted covariance matrix)

Statistical Coupling Analysis (SCA)

Cijab: frequency-based correlation between position i and j

~Cijab is a measure of the significance of observed correlations as judged by the conservation of the amino acids under consideration

After binary approximation:

Binary approximation

Di(ai): the conservation of ai, which is the most prevalent amino acid at that position

Spectral cleaning to separate functional correlation from statistical and historical noisePrincipal Component Analysis

Spectral decomposition of ~Cij matrix to partially sort out the different contributions to the correlations

223 eigenvalues

Lowest 218 – Statistical noiseRandomized alignments retaining the same size and amino acid propensities at sites show eigenvalues of similar magnitude

First mode makes the dominant contribution to ~Cij – historical noise

The first eiganvelue is well approximated by a first order approximation, proves that the first eigenvector should just report the net contribution of each position to the total correlation

Sector Identification using modes 2 to 5

Overview of Sectors

Statistical IndependenceCompute correlation entropy to quantitatively measure the independence of sectors

Minimum discriminatory information method

i.e.

S is small set of position, specifically, the top five positions contributing to each sector

Structure Connectivity

No sector

Known primary/secondary/subdomain-architecture subdivision

Distinction in degree of solvent exposure

Difference in proximity to the active site (not for green sector)

Without information about tertiary structure and only ~10% of total sequence positions contributes strongly to each sector, each sector reveals obvious intra-sector physical connectivity and only a few inter-sector contacts.

Red: focus on S1 pocket

catalytic specificity

Blue: more distributed property

Green: focus around catalytic triad

catalytic activity

Biochemical Independence

Additive effects from combination of mutations between two groups

(magenta: observed | white: predicted)

Mutations of red and blue sectors showed very different effects focused either on catalytic power or thermal stability

Independent Sequence Divergence

Sequence similarity analysis of each sector classifies members in the family effectively only by the related property, while the analysis on all positions failed to do the classification (442 members with functional annotation)

Evidence of “Sector” theory in Other Protein Families

PDZ

PAS

SH2

SH3

Different regulatory mechnisms

Novel Structural Organization

Implication for Physical Properties of Proteins

Alternative View to Calculate Residue Covariance

Technical Challenges

Protein Modulization Adaptive Advantage

Discussion