Covalent modification of an exposed surface turn alters the global conformation of the biotin carrier domain of Escherichia coli acetyl-CoA carboxylase.

摘要: We have studied the apo (unbiotinylated) and holo (biotinylated) forms of BCCP87, an 87-residue COOH-terminal peptide comprising biotin carrier domain carboxyl protein subunit Escherichia coli acetyl-CoA carboxylase. The spontaneously formed disulfide-linked dimers was modified readily by sulfhydryl reagents, whereas remained monomeric unreactive toward reagents unless a denaturant present. These data indicated that single cysteine residue (Cys-116) much more reactive in form protein. Incubation apoBCCP87 with ligase for different times, followed reaction fluorescein-5-maleimide, clearly showed loss Cys-116 reactivity result modification biotin. In addition, 5,5′-dithiobis(2-nitrobenzoic acid) denatured at lower urea concentrations than holoBCCP87. also found least 10-fold sensitive to proteolysis range proteases. Identification cleavage sites differences protease sensitivity could not be attributed shielding susceptible bonds moiety indicate conformational change accompanies biotinylation domain. Thus, β-turn protruding from surface results alteration overall structure this