Protein Data Bank in Europe

Abstract

The class II chelatase CfbA catalyzes Ni₂₊ 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 in vitro. 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 Ni₂₊ and Co₂₊ remains elusive. To understand the structural basis of the underlying mechanisms and evolutionary aspects of the class II chelatases, X-ray crystal structures of Methanocaldococcus jannaschii wild-type CfbA with various ligands, including SHC, Ni₂₊, Ni-SHC, and Co₂₊ were determined. Further, X-ray crystallographic snapshot analysis captured a unique Ni₂₊-SHC-His intermediate complex and Co-SHC-bound CfbA, which resulted from a more rapid chelatase reaction for Co₂₊ than Ni₂₊. Meanwhile, an in vitro activity assay confirmed the different reaction rates for Ni₂₊ and Co₂₊ by CfbA. Based on these structural and functional analyses, the following substrate-SHC-assisted Ni₂₊ insertion catalytic mechanism was proposed: Ni₂₊ insertion to SHC is promoted by the support of an acetate side chain of SHC.