The Heme-Lys Cross-Link in Cytochrome P460 Promotes Catalysis by Enforcing Secondary Coordination Sphere Architecture.

Publication Type:

Journal Article


Biochemistry, Volume 59, Issue 24, p.2289-2298 (2020)


<p>Cytochrome (cyt) P460 is a -type monoheme enzyme found in ammonia-oxidizing bacteria (AOB) and methanotrophs; additionally, genes encoding it have been found in some pathogenic bacteria. Cyt P460 is defined by a unique post-translational modification to the heme macrocycle, where a lysine (Lys) residue covalently attaches to the 13&#39; carbon of the porphyrin, modifying this heme macrocycle into the enzyme&#39;s eponymous P460 cofactor, similar to the cofactor found in the enzyme hydroxylamine oxidoreductase. This cross-link imbues the protein with unique spectroscopic properties, the most obvious of which is the enzyme&#39;s green color in solution. Cyt P460 from the AOB is a homodimeric redox enzyme that produces nitrous oxide (NO) from 2 equiv of hydroxylamine. Mutation of the Lys cross-link results in spectroscopic features that are more similar to those of standard cyt &#39; proteins and renders the enzyme catalytically incompetent for NHOH oxidation. Recently, the necessity of a second-sphere glutamate (Glu) residue for redox catalysis was established; it plausibly serves as proton relay during the first oxidative half of the catalytic cycle. Herein, we report the first crystal structure of a cross-link deficient cyt P460. This structure shows that the positioning of the catalytically essential Glu changes by approximately 0.8 Å when compared to a cross-linked, catalytically competent cyt P460. It appears that the heme-Lys cross-link affects the relative position of the P460 cofactor with respect to the second-sphere Glu residue, therefore dictating the catalytic competency of the enzyme.</p>