Novel Bcl-2 homology-3 domain-like sequences identified from screening randomized peptide libraries for inhibitors of the pro-survival Bcl-2 proteins.
Interactions between Bcl-2 homology-3 (BH3)-only proteins and their pro-survival Bcl-2 family binding partners initiate the intrinsic apoptosis pathway. These interactions are mediated by a short helical motif, the BH3 domain, on the BH3-only protein, which inserts into a hydrophobic groove on the pro-survival molecule. To identify novel peptidic ligands that bind Mcl-1, a pro-survival protein relative of Bcl-2, both human and mouse Mcl-1 were screened against large randomized phage-displayed peptide libraries. We identified a number of 16-mer peptides with sub-micromolar affinity that were highly selective for Mcl-1, as well as being somewhat selective for the species of Mcl-1 (human or mouse) against which the library was panned. Interestingly, these sequences all strongly resembled natural BH3 domain sequences. By switching residues within the best of the human Mcl-1-binding sequences, or extending beyond the core sequence identified, we were able to alter the pro-survival protein interaction profile of this peptide such that it now bound all members tightly and was a potent killer when introduced into cells. Introduction of an amide lock constraint within this sequence also increased its helicity and binding to pro-survival proteins. These data provide new insights into the determinants of BH3 domain:pro-survival protein affinity and selectivity.
Lee
Erinna F
EF
Walter and Eliza Hall Institute of Medical Research, 1G Royal Pde., Parkville, Victoria 3052, Australia.
Fedorova
Anna
A
Zobel
Kerry
K
Boyle
Michelle J
MJ
Yang
Hong
H
Perugini
Matthew A
MA
Colman
Peter M
PM
Huang
David C S
DC
Deshayes
Kurt
K
Fairlie
W Douglas
WD
D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D064549:Myeloid Cell Leukemia Sequence 1 Protein; D019151:Peptide Library; D010455:Peptides / Q000138:chemical synthesis / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity
C579094:Mcl1 protein, mouse; D064549:Myeloid Cell Leukemia Sequence 1 Protein; D019151:Peptide Library; D010455:Peptides; D019253:Proto-Oncogene Proteins c-bcl-2