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Updated: Jun 17, 2026

Biotinylated Cell-penetrating Peptides to Study Intracellular Protein-protein Interactions
Published on: December 20, 2017
The polypeptide Syn67 interacts physically with human holocarboxylase synthetase, but is not a target for
Yousef I Hassan1, Hideaki Moriyama, Janos Zempleni
1Department of Nutrition and Health Sciences, University of Nebraska at Lincoln, NE 68583-0806, USA.
None:
Holocarboxylase synthetase (HCS) catalyzes the binding of biotin to lysines in carboxylases and histones in two steps. First, HCS catalyzes the synthesis of biotinyl-5'-AMP; second, the biotinyl moiety is ligated to lysine residues. It has been proposed that step two is fairly promiscuous, and that protein biotinylation may occur in the absence of HCS as long as sufficient exogenous biotinyl-5'-AMP is provided. Here, we identified a novel polypeptide (Syn67) with a basic patch of lysines and arginines. Yeast-two-hybrid assays and limited proteolysis assays revealed that both N- and C-termini of HCS interact with Syn67. A potential target lysine in Syn67 was biotinylated by HCS only after arginine-to-glycine substitutions in Syn67 produced a histone-like peptide. We identified a Syn67 docking site near the active pocket of HCS by in silico modeling and site-directed mutagenesis. Biotinylation of proteins by HCS is more specific than previously assumed.
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