Structural Evidence for DUF512 as a Radical <i>S</i>-Adenosylmethionine Cobalamin-Binding Domain
Bo Wang, Amy E. Solinski, Matthew I. Radle, Olivia M. Peduzzi, Hayley Knox, Jinhe Cui, Ravi K. Maurya, Neela H. Yennawar, Squire J. Booker
Abstract
High Resolution Image Download MS PowerPoint Slide Cobalamin (Cbl)-dependent radical S -adenosylmethionine (SAM) enzymes constitute a large subclass of radical SAM (RS) enzymes that use Cbl to catalyze various types of reactions, the most common of which are methylations. Most Cbl-dependent RS enzymes contain an N-terminal Rossmann fold that aids Cbl binding. Recently, it has been demonstrated that the methanogenesis marker protein 10 (Mmp10) requires Cbl to methylate an arginine residue in the α-subunit of methyl coenzyme M reductase. However, Mmp10 contains a Cbl-binding domain in the C-terminal region of its primary structure that does not share significant sequence similarity with canonical RS Cbl-binding domains. Bioinformatic analysis of Mmp10 identified DUF512 (Domain of Unknown Function 512) as a potential Cbl-binding domain in RS enzymes. In this paper, four randomly selected DUF512-containing proteins from various organisms were overexpressed, purified, and shown to bind Cbl. X-ray crystal structures of DUF512-containing proteins from Clostridium sporogenes and Pyrococcus furiosus were determined, confirming their C-terminal Cbl-binding domains. The structure of the DUF512-containing protein from C. sporogenes is the first of an RS enzyme containing a PDZ domain. Its RS domain has an unprecedented β 3 α 4 core, whereas most RS enzymes adopt a (βα) 6 core. The DUF512-containing protein from P. furiosus has no PDZ domain, but its RS domain also has an uncommon (βα) 5 core.