COMPUTING OF THE RAD51 -BRCA2 INTERACTIONS Assist. Prof. Albena Todorova, PhD Andrey Kirov, Msc Peycho Petkov, PhD Computing of the effects of genetic.

COMPUTING OF THE RAD51 -BRCA2

INTERACTIONS

Assist. Prof. Albena Todorova, PhD

Andrey Kirov, Msc

Peycho Petkov, PhD

Computing of the effects of genetic mutations in the BRCA2 gene to the protein structure and evaluation of these

changes to their interactions with other proteins

WHY?

519 000 women died in 2004 due to breast cancer, and although breast cancer is thought to be a disease of the developed world, a majority (69%) of all breast cancer deaths occurs in developing countries (WHO Global Burden of Disease, 2004)

According to WHO breast cancer is the most common cancer in women worldwide, comprising 16% of all female cancers

Breast cancer survival rates vary greatly worldwide, ranging from 80% or over in North America, Sweden and Japan to around 60% in middle-income countries and below 40% in low-income countries (Coleman et al., 2008). The low survival rates in less developed countries can be explained mainly by the lack of early detection and prophylactics programmes

Family history of breast or ovarian cancer - increases the risk by a factor of two or three. Some mutations, particularly in BRCA1 and BRCA2 genes result in a very high risk for breast cancer

RISK FACTORS

Age – cancer risk increases with aging, no matter of genetic predispositions

Hormone influence – high estrogen level increases the risk

Obesity

Nutrition diete

DNA ISOLATION

BRCA GENETIC TESTS ALGORITHM

SEQUENCING OF BRCA1 & BRCA2 GENES

qPCR/MLPA ANALYSIS OF BRCA1 & BRCA2 GENES

NORMAL RESULT

EVERYTHING IS OK

LARGE DELETIONS/DUPLICATIONS

ARE DETECTED

INCRESED RISK OF BREAST AND

OVARIAN CANCER

MUTATION WITH KNOWN

GENOTYPE-PHENOTYPE

CORRELATION

DETECTION OF POINT MUTATIONS

MUTATION WITH UNKNOWN GENOTYPE-PHENOTYPE

CORRELATION

?

BRCA2

The BRCA2 tumour suppressor is essential for homologous DNA recombination, a process for the errorfree repair of DNA double-stranded breaks , in the cells of higher eukaryotes

BRCA2 central function during recombination is to control the RAD51 recombinase, which is indispensable for DNA recombination

The homologous DNA recombination begins when broken DNA is end-resected to generate ssDNA, upon which RAD51 oligomerizes to form an active nucleoprotein filament. The RAD51 nucleoprotein filament catalyzes strand exchange by invading and pairing with a homologous DNA duplex, used as a template for repair

Reymera et al., 2009

BRCA2-RAD51 INTERACTIONS

Two distinct regions in human BRCA2 bind directly to RAD51:

1.Eight BRC repeats, evolutionarily conserved motifs of approximately 35 residues each, distributed in a 1100 residue region of BRCA2 (BRC1-8) - conserved amongst vertebrate orthologues in both their sequence and spacing

2. C-terminus RAD51-interacting region - stabilizes RAD51 oligomericassemblies in vitro both on and off DNA

BRCA2-RAD51 INTERACTIONS

BRCA2 BRC repeats

SIMULATION OF BRCA2-RAD51 INTERACTIONS

PROBLEMS The whole 3D structure of BRCA2 is still unknown, we have information for the 3D structure of region of BRC4 repeat:

Aminoacid sequence of BRC4 repeat: PTLLGFHTASGKKVKIAKESLDKVKNLFDEKEQ

Position: 1524 1546

Crystal structure of a RAD51-BRCA2 BRC4 repeat complex

DOI:10.2210/pdb1n0w/pdb

Source: RCSB Protein Data Bank

SIMULATION OF BRCA2-RAD51 INTERACTIONS

PROBLEMS In this region we have very limited information about the correlation

Genotype - Phenotype

Solution

In this region we have 1 mutation with known correlation Genotype-Phenotype and used it to test our model

Computing the interaction of the normal/wild type BRC4 and RAD51 and its use as a base to compare

Mutation №

Mutation SR-LJ Coul-SR SR-Energy

1 Wild type -322.345 -376.139 -698.4842 Trp1520Ala -381.386 -409.69 -791.0763 Phe1524Val -325.782 -319.335 -645.1174 Gly1529Arg -365.424 -341.879 -707.3035 Pro1519His -306.214 -307.031 -613.2456 Pro1519Lys -377.373 -411.534 -788.9077 Leu1521Ser -301.457 -264.946 -566.4038 Phe1524Tyr -336.86 -359.267 -696.1279 Phe1524Trp -256.996 -339.593 -596.589

10 Phe1524Gly -260.885 -220.265 -481.1511 Phe1546Tyr -469.893 -494.114 -964.00712 Phe1546Trp -305.869 -294.978 -600.84713 Phe1546Ser -331.077 -355.703 -686.7814 Asp1547Tyr -360.436 -348.169 -708.605

SIMULATION OF BRCA2-RAD51 INTERACTIONS

Normal Phenotype

CONCLUSION

THE STRUCTURAL HOMOLOGY IN THE REGION OF THE TWO HIDROPHOBIC POCKETS IS MORE IMPORTANT FOR THE BRC4-

RAD51 COMPLEX STABILITY THAN THE HIDROPHOBIC INTERACTIONS INSIDE THE POCKET

CONTROVERSARY

Position: 1524 1546A peptide containing both F1524 and F1546 mutated to tryptophan retains similar

behaviour

SO?WHAT TO

DO?

MORE RESEARCH

Acknowledgement:

Financial support of Bulgarian Supercomputing Centre (BGSC) is greatly

appreciated – Grant 10/2009