Mechanistic significance of the preparatory migration of hydrogen atoms around the FeMo-co active site of nitrogenase

The migration of H atoms over S and Fe atoms in the reaction domain of FeMo-co, the active site of nitrogenase, is described and used to explain mechanistic data on the catalyzed reductions of N-2 and C2H2. After electron transfer to FeMo-co, H atoms are generated by fast proton supply to S3B (atom labels from structure 1M1N) and migrate vectorially via several pathways from S3B to locations on the FeMo-co face, specifically Fe6, S2B, Fe2, and S2A (calculated reaction profiles are reported). The EnHn reduction levels (n = 1-4) in the Thorneley-Lowe kinetic-mechanistic schemes are each potential sequences of substructures with different distributions of H atoms. The positions of H atoms influence the binding of substrates N-2 and C2H2, and the bound substrate subsequently blocks further migration of H atoms past the binding site. This model provides a consistent structural interpretation of (a) the two-site reactivity of C2H2 and the differentiation of the high- and low-affinity sites as due to different preparatory H migration; (b) the differing mutual inhibitions of N-2 and C2H2 in wild-type protein; (c) the modified reactivity of the Azotobacter vinelandii alpha-(Gly)69(Ser) mutant with N-2 and C2H2; and (d) the basis for the stereoselectivity of hydrogenation of C2D2 and its loss in some mutant proteins. Some structures for initially bound N-2 and C2H2, and their hydrogenated intermediates, are presented. The key new concept is that binding sites and binding states for substrates and intermediates are characterized not only by their locations on the FeMo-co face but also by the structural and temporal status of the distribution of H atoms over the FeMo-co reaction domain.