discovery of the myxosortases that process MYXO-CTERM and three novel prokaryotic C-terminal protein-sorting signals that share invariant Cys residues.

The LPXTG protein-sorting signal, found in surface proteins of various Gram-positive pathogens, was the founding member of a growing panel of prokaryotic small C-terminal sorting domains. Sortase A cleaves LPXTG, exosortases (XrtA and XrtB) cleave the PEP-CTERM sorting signal, archaeosortase A cleaves PGF-CTERM, and rhombosortase cleaves GlyGly-CTERM domains. Four sorting signal domains without previously known processing proteases are the MYXO-CTERM, JDVT-CTERM, Synerg-CTERM, and CGP-CTERM domains. These exhibit the standard tripartite architecture of a short signature motif, a hydrophobic transmembrane segment, and an Arg-rich cluster. Each has an invariant cysteine in its signature motif. Computational evidence strongly suggests that each of these four Cys-containing sorting signals is processed, at least in part, by a cognate family of glutamic-type intramembrane endopeptidases related to the eukaryotic type II CAAX-processing protease Rce1. For the MYXO-CTERM sorting signals of different lineages, their sorting enzymes, called myxosortases, include MrtX (MXAN_2755 in ), MrtC, and MrtP, all with radically different N-terminal domains but with a conserved core. Related predicted sorting enzymes were also identified for JDVT-CTERM (MrtJ), Synerg-CTERM (MrtS), and CGP-CTERM (MrtA). This work establishes a major new family of protein-sorting housekeeping endopeptidases contributing to the surface attachment of proteins in prokaryotes. IMPORTANCE Homologs of the eukaryotic type II CAAX-box protease Rce1, a membrane-embedded endopeptidase found in yeast and human ER and involved in sorting proteins to their proper cellular locations, are abundant in prokaryotes but not well understood there. This bioinformatics paper identifies several subgroups of the family as cognate endopeptidases for four protein-sorting signals processed by previously unknown machinery. Sorting signals with newly identified processing enzymes include three novel ones, but also MYXO-CTERM, which had been the focus of previous experimental work in the model fruiting and gliding bacterium . The new findings will substantially improve our understanding of Cys-containing C-terminal protein-sorting signals and of protein trafficking generally in bacteria and archaea.