Peptide Ligands for Pro-survival Protein Bfl‑1 from Computationally Guided Library Screening

• Published in: ACS chemical biology Vol. 8; no. 4; pp. 778 - 788
• Main Authors: Dutta, Sanjib, Chen, T. Scott, Keating, Amy E
• Format: Journal Article
• Language: English
• Published: United States American Chemical Society 19.04.2013
• Subjects: Amino Acid Sequence; Ligands; Minor Histocompatibility Antigens; Models, Molecular; Molecular Sequence Data; Peptide Library; Proto-Oncogene Proteins c-bcl-2 - chemistry; Proto-Oncogene Proteins c-bcl-2 - metabolism
• ISSN: 1554-8929; 1554-8937
• DOI: 10.1021/cb300679a

Pro-survival members of the Bcl-2 protein family inhibit cell death by binding short helical BH3 motifs in pro-apoptotic proteins. Mammalian pro-survival proteins Bcl-xL, Bcl-2, Bcl-w, Mcl-1, and Bfl-1 bind with varying affinities and specificities to native BH3 motifs, engineered peptides, and small molecules. Biophysical studies have determined interaction patterns for these proteins, particularly for the most-studied family members Bcl-xL and Mcl-1. Bfl-1 is a pro-survival protein implicated in preventing apoptosis in leukemia, lymphoma, and melanoma. Although Bfl-1 is a promising therapeutic target, relatively little is known about its binding preferences. We explored the binding of Bfl-1 to BH3-like peptides by screening a peptide library that was designed to sample a high degree of relevant sequence diversity. Screening using yeast-surface display led to several novel high-affinity Bfl-1 binders and to thousands of putative binders identified through deep sequencing. Further screening for specificity led to identification of a peptide that bound to Bfl-1 with K d < 1 nM and very slow dissociation from Bfl-1 compared to other pro-survival Bcl-2 family members. A point mutation in this sequence gave a peptide with ∼50 nM affinity for Bfl-1 that was selective for Bfl-1 in equilibrium binding assays. Analysis of engineered Bfl-1 binders deepens our understanding of how the binding profiles of pro-survival proteins differ and may guide the development of targeted Bfl-1 inhibitors.