Modeling and numerical simulation of biotin carboxylase kinetics: implications for half-sites reactivity.
Biotin carboxylase catalyzes the ATP-dependent carboxylation of biotin and is one component of the multienzyme complex acetyl-CoA carboxylase that catalyzes the first committed step in fatty acid synthesis in all organisms. In Escherichia coli, biotin carboxylase exists as a homodimer where each subunit contains a complete active site. In a previous study (Janiyani, K., Bordelon, T., Waldrop, G.L., Cronan Jr., J.E., 2001. J. Biol. Chem. 276, 29864-29870), hybrid dimers were constructed where one subunit was wild-type and the other contained an active site mutation that reduced activity at least 100-fold. The activity of the hybrid dimers was only slightly greater than the activity of the mutant homodimers and far less than the expected 50% activity for completely independent active sites. Thus, there is communication between the two subunits of biotin carboxylase. The dominant negative effect of the mutations on the wild-type active site was interpreted as alternating catalytic cycles of the active sites in the homodimer. In order to test the hypothesis of oscillating catalytic cycles, mathematical modeling and numerical simulations of the kinetics of wild-type, hybrid dimers, and mutant homodimers of biotin carboxylase were performed. Numerical simulations of biotin carboxylase kinetics were the most similar to the experimental data when an oscillating active site model was used. In contrast, alternative models where the active sites were independent did not agree with the experimental data. Thus, the numerical simulations of the proposed kinetic model support the hypothesis that the two active sites of biotin carboxylase alternate their catalytic cycles.
de Queiroz
Marcio S
MS
Department of Mechanical Engineering, Louisiana State University, Baton Rouge, LA 70803-6413, USA. dequeiroz@me.lsu.edu
Waldrop
Grover L
GL
D000255:Adenosine Triphosphate / Q000378:metabolism*; D000818:Animals; D001665:Binding Sites / Q000235:genetics; D001710:Biotin / Q000378:metabolism*; D019731:Carbon-Nitrogen Ligases / Q000235:genetics / Q000378:metabolism*; D003198:Computer Simulation; D019281:Dimerization; D008956:Models, Chemical; D009154:Mutation
D001710:Biotin; D000255:Adenosine Triphosphate; D019731:Carbon-Nitrogen Ligases; C019791:biotin carboxylase