Expression in Escherichia coli of N- and C-terminally Deleted Human Holocarboxylase Synthetase INFLUENCE OF THE N-TERMINUS ON BIOTINYLATION AND IDENTIFICATION OF A MINIMUM FUNCTIONAL PROTEIN

摘要: Biotin functions as a covalently bound cofactor of biotindependent carboxylases. attachment is catalyzed by biotin protein ligases, called holocarboxylase synthetase in mammals and BirA prokaryotes. These enzymes show high degree sequence similarity their biotinylation domains but differ markedly the length N terminus. also repressor operon, its DNA site located The function eukaryotic terminus unknown. Holocarboxylase with N- C-terminal deletions were evaluated for ability to catalyze after expression Escherichia coli using bacterial human acceptor substrates. We showed that minimum functional comprised last 349 726-residue protein, which includes domain. Significantly, enzyme containing intermediate length, N-terminal interfered transfer interaction different peptide propose contributes through interactions may affect substrate recognition. Our findings provide rationale responsiveness patients point mutations synthetase. Such mutant respond biotin-mediated stabilization substrate-bound complex.