6c4u

Publication Abstract from PubMed

Transcription factor c-Myc is an oncoprotein that is regulated at the post-translational level through phosphorylation of two conserved residues, Serine 62 (Ser62) and Threonine 58 (Thr58). A highly specific tool capable of recognizing Myc via pThr58 is needed to monitor activation and localization. Through phage display, we have isolated 10 engineered Forkhead-associated (FHA) domains that selectively bind to a phosphothreonine (pThr)-containing peptide (53-FELLPpTPPLSPS-64) segment of human c-Myc. One domain variant was observed to bind to the Myc-pThr58 peptide with a KD value of 800nM and had >1,000-fold discrimination between the phosphorylated and non-phosphorylated peptide. The crystal structure of the engineered FHA Myc-pThr-binding domain (Myc-pTBD) was solved in complex with its cognate ligand. The Myc-pTBD was observed to be structurally similar to the yeast Rad9 FHA1 domain, except that its beta4-beta5 and beta10-beta11 loops form a hydrophobic pocket to facilitate the interaction between the domain and the peptide ligand. The Myc-pTBD's specificity for its cognate ligand was demonstrated to be on a par with 3 commercial polyclonal antibodies, suggesting that this recombinant reagent is a viable alternative to antibodies for monitoring Myc regulation.

Generating a recombinant phosphothreonine-binding domain for a phosphopeptide of the human transcription factor, c-Myc.,Venegas LA, Kall SL, Bankole O, Lavie A, Kay B N Biotechnol. 2018 May 12. pii: S1871-6784(18)30120-1. doi:, 10.1016/j.nbt.2018.05.001. PMID:29763736^[1]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.