A mixed-valent Fe(II)Fe(III) species converts cysteine to an oxazolone/thioamide pair in methanobactin biosynthesis

Abstract

Significance Methanobactins (Mbns), copper-binding peptidic compounds produced by some bacteria, are candidate therapeutics for human diseases of copper overload. The paired oxazolone-thioamide bidentate ligands of methanobactins are generated from cysteine residues in a precursor peptide, MbnA, by the MbnBC enzyme complex. MbnBC activity depends on the presence of iron and oxygen, but the catalytically active form has not been identified. Here, we provide evidence that a dinuclear Fe(II)Fe(III) center in MbnB, which is the only representative of a >13,000-member protein family to be characterized, is responsible for this reaction. These findings expand the known roles of diiron enzymes in biology and set the stage for mechanistic understanding, and ultimately engineering, of the MbnBC biosynthetic complex.

A remarkable transformation catalyzed by a domain-of-unknown-function 692 during the biosynthesis of a new RiPP natural product

“…Formation of a similar β-lactam was also proposed for MbnBC catalysis as well as other mononuclear nonheme iron enzymes such as isopenicillin N synthase. 20,22,[47][48][49] At this point, the reaction of ChrHI diverges from previous enzymes in that a second nucleophilic attack of the amide nitrogen of the downstream Met10 onto the carbonyl of the β-lactam would result in the formation of a bicyclic intermediate III. For MbnBC, the -lactam is attacked by the oxygen atom of the upstream amide, 21 possibly with the intermediacy of an enzyme bound intermediate.…”

Unusual Dioxygen-Dependent Reactions Catalyzed by Nonheme Iron Enzymes in Natural Product Biosynthesis

“…MbnC is responsible for the recognition of the substrate MbnA, and MbnB and MbnC form a heterotrimeric complex with MbnA. Initial investigations of the purified MbnBC complex by Mössbauer spectroscopy and iron quantification suggested that it contained di- or trinuclear clusters . MbnBC reacts with MbnA to install the oxazolone–thioamide group in 245 in the presence of O 2 .…”

“…Initial investigations of the purified MbnBC complex by Mossbauer spectroscopy and iron quantification suggested that it contained di-or trinuclear clusters. 253 MbnBC reacts with MbnA to install the oxazolone− thioamide group in 245 in the presence of O 2 . This unusual fourelectron oxidation reaction may be coupled with the fourelectron reduction of O 2 because no external reducing cosubstrate is needed.…”

“…254 Although little is known about the mechanism of MbnABC, a possible mechanism has been proposed (Scheme 27). 253,254 In Some enzyme residues (such as Asp240) could be involved in this oxazolone-forming step. 254 However, the exact mechanism of this complex transformation remains to be investigated.…”

“…Although little is known about the mechanism of MbnABC, a possible mechanism has been proposed (Scheme ). , In the mechanisms of mixed-valent diiron oxygenases and oxidases (MVDOs) such as myo -inositol oxygenase (MIOX) and PhnZ, the substrate and O 2 cosubstrate coordinate the Fe­(III) and Fe­(II) centers, respectively. Thus, MbnBC would also bind the thiol of 239 and O 2 substrates in a similar manner.…”