A novel catalytic heme cofactor in SfmD with a single thioether bond and a <i>bis</i>-His ligand set revealed by a <i>de novo</i> crystal structural and spectroscopic study
Inchul Shin, Ian Davis, Karinel Nieves-Merced, Yifan Wang, Stanton F. McHardy, Aimin Liu
Abstract
, His274 and His313. We demonstrated that SfmD heme is catalytically active and can utilize dioxygen and ascorbate for a single-oxygen insertion into 3-methyl-l-tyrosine. Catalytic assays using ascorbate derivatives revealed the functional groups of ascorbate essential to its function as a cosubstrate. Abolishing the thioether linkage through mutation of Cys317 resulted in catalytically inactive SfmD variants. EPR and optical data revealed that the heme center undergoes a substantial conformational change with one axial histidine ligand dissociating from the iron ion in response to substrate 3-methyl-l-tyrosine binding or chemical reduction by a reducing agent, such as the cosubstrate ascorbate. The labile axial ligand was identified as His274 through redox-linked structural determinations. Together, identifying an unusual heme cofactor with a previously unknown heme-binding motif for a monooxygenase activity and the structural similarity of SfmD to the members of the heme-based tryptophan dioxygenase superfamily will broaden understanding of heme chemistry.