The nickel-sirohydrochlorin formation mechanism of the ancestral class II chelatase CfbA in coenzyme F430 biosynthesis.

The class II chelatase CfbA catalyzes Ni insertion into sirohydrochlorin (SHC) to yield the product nickel-sirohydrochlorin (Ni-SHC) during coenzyme F430 biosynthesis. CfbA is an important ancestor of all the class II chelatase family of enzymes, including SirB and CbiK/CbiX, functioning not only as a nickel-chelatase, but also as a cobalt-chelatase . Thus, CfbA is a key enzyme in terms of diversity and evolution of the chelatases catalyzing formation of metal-SHC-type of cofactors. However, the reaction mechanism of CfbA with Ni and Co remains elusive. To understand the structural basis of the underlying mechanisms and evolutionary aspects of the class II chelatases, X-ray crystal structures of wild-type CfbA with various ligands, including SHC, Ni, Ni-SHC, and Co were determined. Further, X-ray crystallographic snapshot analysis captured a unique Ni-SHC-His intermediate complex and Co-SHC-bound CfbA, which resulted from a more rapid chelatase reaction for Co than Ni. Meanwhile, an activity assay confirmed the different reaction rates for Ni and Co by CfbA. Based on these structural and functional analyses, the following substrate-SHC-assisted Ni insertion catalytic mechanism was proposed: Ni insertion to SHC is promoted by the support of an acetate side chain of SHC.