The Cytochrome P450 Catalyzing C−S Bond Formation in S‐Heterocyclization of Chuangxinmycin Biosynthesis

Abstract

Microbial sulfur-containing secondary metabolites show various biological activities,but the CÀSbond-forming in their biosynthetic metabolism has not been thoroughly understood. Here,w ep resent genetic, biochemical and structural characterization of acytochrome P450 monooxygenase CxnD exhibiting C À Sbond forming activity in S-heterocyclization of chuangxinmycin biosynthesis.I nv ivo and in vitro analyses demonstrated that CxnD generated an indole-fused dihydrothiopyran skeleton from a l-Trp-derived thiol intermediate. Furthermore,X-ray crystal structure of CxnD in complex with asubstrate analogue and structure-based mutagenesis revealed intimate details of the substrate binding mode.Aradical mechanism initiated by abstraction of the imino hydrogen atom or an electron from indole group of the substrate was proposed for CxnD,whichprovided valuable insights into the molecular basis for the intra-molecular C(sp 2 )ÀHthiolation by the P450 in chuangxinmycin biosynthesis.

Biosynthesis of Chuangxinmycin Featuring a Deubiquitinase‐like Sulfurtransferase

“…It should be noted that overlapping in time frame with our work, Hong and coworkers reported the same function of CxnD (a homolog of Cxm5). [23] Based upon the established individual biosynthetic steps for TPI skeleton assembly, we sought to reconstitute the biosynthetic pathway from 2 to 10 in vitro. Particularly, when 2 was firstly incubated with an equal concentration (10 mM) of Cxm3, Cxm4, CxmJ, CxmM, Cxm5, Cxm6 and Cxm7 for 6 h in the presence of co-factors, 2 was mostly consumed, and the intermediate 3 was also efficiently converted into 4; while 4 was only partially transformed to 10 (Figure 4 B, trace i, ii).…”

Role of “S” Substitution on C–H Activation Reactivity of Iron(IV)–Oxo Cyclam Complexes: a Computational Investigation

“…There are several sulfur-ligated Fe­(IV)O centers also being found as key oxidants in the catalytic cycles of various heme and non-heme iron oxygenases, for example, cysteine dioxygenase (CDO) that catalyzes the transformation of l -cysteine to cysteine sulfinic acid, ergothioneine biosynthesis enzymes (OvoA), and so forth. Ligation of sulfur atoms has been reported several times as they are suggested to be reactive intermediates for a wide range of chemical transformations including oxygenation, hydroxylation, and C–S bond formation reactions . Fe­(IV)O porphyrin p-cation radical (Cpd I) intermediates containing a thiolate ligand trans to the oxo group have been characterized in a number of heme enzymes .…”

“…Ligation of sulfur atoms has been reported several times as they are suggested to be reactive intermediates for a wide range of chemical transformations including oxygenation, 36 hydroxylation, 37 and C−S bond formation reactions. 38 Fe(IV)�O porphyrin p-cation radical (Cpd I) intermediates containing a thiolate ligand trans to the oxo group have been characterized in a number of heme enzymes. 37 A lot of investigations have also been presented related to the replication of CDO active sites toward synthetic analogues.…”

Cytochrome P450s in algae: Bioactive natural product biosynthesis and light-driven bioproduction

“…During the biosynthesis of natural products, generally, P450s implement their functions by catalyzing the common monooxygenation ( e.g ., hydroxylation and epoxidation) reactions to decorate the core structure, improve the aqueous solubility of products, and provide chemical handles for further modifications 15 . Intriguingly, in an increasing number of cases, P450s have been found capable of mediating diverse “uncommon” reactions for skeleton construction, including C–X (X = C/N/S) bond formation/scission, ring formation/expansion/contraction and even more vagarious reactions, which dramatically expand the chemical space of natural products 16 , 17 , 18 , 19 , 20 . As a result, the P450-mediated reactions greatly increase the diversity of both the chemical structures and biological activities of natural products.…”