Structural basis of phosphopeptide recognition by the BRCT domain of BRCA1

Williams, R. Scott; Lee, Megan S.; Hau, David; Glover, J. N. Mark

doi:10.1038/nsmb776

Cited by 221 publications

(313 citation statements)

References 35 publications

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“…Structural and biochemical studies between BRCT(BRCA1) and phosphorylated peptides have lead to the identification of a pSXXF as the binding motif on the peptide and mapped the binding site to a region at the interface of the two BRCT domains of BRCA1 [12,14,[18][19][20][21][22][23][24]. The BRCT-phosphoprotein interactions are transient and structural studies show that BACH1 and CtIP bind to the same site on the BRCT domains.…”

Section: Introductionmentioning

confidence: 99%

Dual-fluorophore quantitative high-throughput screen for inhibitors of BRCT–phosphoprotein interaction

Simeonov

Yasgar

Jadhav

et al. 2008

Analytical Biochemistry

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Finding specific small-molecule inhibitors of protein-protein interactions remains a significant challenge. Recently, attention has grown toward "hot-spot" interactions where binding is dominated by a limited number of amino acid contacts, theoretically offering an increased opportunity for disruption by small molecules. Inhibitors of the interaction between BRCT (Cterminal portion of BRCA1, a key tumor suppressor protein with various functions), and phosphorylated protein (Abraxas, BACH1, CtIP) implicated in DNA damage response and repair pathways, should prove useful in studies of BRCA1's role in cancer and to potentially sensitize tumors to chemotherapeutic agents. We developed and miniaturized to 1536-well format and 3 μL final volume a pair of fluorescence polarization (FP) assays utilizing fluorescein-and rhodaminelabeled pBACH1 fragment. In order to minimize the effect of fluorescence artifacts and to increase the overall robustness of the screen, the 75,552 compound library members were each assayed against both the fluorescein-and rhodamine-labeled probe-protein complexes in separate but interleaved reactions. In addition, every library compound was tested over a range of concentrations, following the qHTS paradigm (Inglese et al, PNAS, 103, 1147(2006). Analyses of the screening results led to the selection and subsequent confirmation of 16 compounds active in both assays. Faced with a traditionally difficult protein-protein interaction assay, by performing two-fluorophore qHTS we were able to confidently select a number of actives for further studies.

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Section: Introductionmentioning

confidence: 99%

Dual-fluorophore quantitative high-throughput screen for inhibitors of BRCT–phosphoprotein interaction

Simeonov

Yasgar

Jadhav

et al. 2008

Analytical Biochemistry

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“…In the wtBRCA1, the loop in which T1691 is located forms the protein surface and is a part of the binding pocket interacting with the BACH1 phosphorylated peptide 26,27 (Figure 2b). The buried T1691 residue makes the hydrogen bond to S1715 and D1692, as a contribution to hold the structure of protein surface and the binding pocket ( Figure 2c).…”

Section: Bioinformatics Studymentioning

confidence: 99%

Multimodel assessment of BRCA1 mutations in Taiwanese (ethnic Chinese) women with early-onset, bilateral or familial breast cancer

et al. 2012

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Although evidence suggests an importance of genetic factors in the development of breast cancer in Taiwanese (ethnic Chinese) women, including a high incidence of early-onset and secondary contralateral breast cancer, a major breast cancer predisposition gene, BRCA1, has not been well studied in this population. In fact, the carcinogenic impacts of many genetic variants of BRCA1 are unknown and classified as variants of uncertain significance (VUS). It is therefore important to establish a method to characterize the BRCA1 VUSs and understand their role in Taiwanese breast cancer patients. Accordingly, we developed a multimodel assessment strategy consisting of a prescreening portion and a validated functional assay to study breast cancer patients with early-onset, bilateral or familial breast cancer. We found germ-line BRCA1 mutations in 11.1% of our cohort and identified one novel missense mutation, c.5191C4A. Two genetic variants were initially classified as VUSs (c.1155C4T and c.5191C4A). c.1155C4T is not predicted to be deleterious in the prescreening portion of our assessment strategy. c.5191C4A, on the other hand, causes p.T1691K, which is predicted to have high deleterious probability because of significant structural alteration, a high deleterious score in the predictive programs and, clinically, triple negative characteristics in breast tumors. This mutant is confirmed by transcription activation and yeast growth-inhibition assays. In conclusion, we show as high a prevalence of germ-line BRCA1 mutation in high-risk Taiwanese patients as in Caucasians and demonstrate a useful strategy for studying BRCA1 VUSs. Keywords: BRCA1; breast cancer; structure modeling; variants of uncertain significance INTRODUCTION Breast cancer is one of the most common cancers and is the leading cause of cancer-related death in women throughout the world. Interestingly, the prevalence of breast cancer is lower in Taiwanese (ethnic Chinese) populations than in Caucasian populations, 1 and the epidemiological patterns are different in the two patient groups. First, the median age of breast cancer patients in Taiwan (45-49 years) is lesser than that in Western countries (70-74 years). 2 Second, a rise in incidence has been observed consistently in Asian countries, 2-4 and one age-period-cohort analysis showed that the increase in breast cancer incidence was largely due to a rise in young patients. 2 Third, a longitudinal cohort revealed that young breast cancer patients in

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“…Tandem BRCT repeats use the topological switch point-termed the P1 pocket-of their first BRCT domain to bind phospho-serine (pSer) residues in their interaction partners ( Figure 5). Similar to FHA domains, this interaction involves the side chain of a conserved serine residue, but in BRCT domains the phosphate moiety of the pSer residue is further stabilized by the interaction with the side chain of a conserved lysine, as well as the backbone atoms from the glycine immediately following the conserved serine [68].…”

Section: Tandem Brct Domains: the Two-knob Hypothesismentioning

confidence: 99%