Structure of dimerized assimilatory NADPH-dependent sulfite reductase reveals the minimal interface for diflavin reductase binding

Abstract

Escherichia coli NADPH-dependent assimilatory sulfite reductase (SiR) fixes sulfur for incorporation into sulfur-containing biomolecules. SiR is composed of two subunits: an NADPH, FMN, and FAD-binding diflavin reductase and an iron siroheme/Fe₄S₄ cluster-containing oxidase. How they interact has been unknown for over 50 years because SiR is highly flexible, thus has been intransigent for traditional X-ray or cryo-EM structural analysis. A combination of the chameleon plunging system with a fluorinated lipid overcame the challenge of preserving a dimer between the subunits for high-resolution (2.84 Å) cryo-EM analysis. Here, we report the first structure of the reductase/oxidase complex, revealing how they interact in a minimal interface. Further, we determined the structural elements that discriminate between pairing a siroheme-containing oxidase with a diflavin reductase or a ferredoxin partner to channel the six electrons that reduce sulfite to sulfide.