title: Assembly and function of a spore coat-associated transglutaminase of Bacillus subtilis.
abstract: The assembly of a multiprotein coat around the Bacillus subtilis spore confers resistance to lytic enzymes and noxious chemicals and ensures normal germination. Part of the coat is cross-linked and resistant to solubilization. The coat contains epsilon-(gamma-glutamyl)lysyl cross-links, and the expression of the gene (tgl) for a spore-associated transglutaminase was shown before to be required for the cross-linking of coat protein GerQ. Here, we have investigated the assembly and function of Tgl. We found that Tgl associates, albeit at somewhat reduced levels, with the coats of mutants that are unable to assemble the outer coat (cotE), that are missing the inner coat and with a greatly altered outer coat (gerE), or that are lacking discernible inner and outer coat structures (cotE gerE double mutant). This suggests that Tgl is present at various levels within the coat lattice. The assembly of Tgl occurs independently of its own activity, as a single amino acid substitution of a cysteine to an alanine (C116A) at the active site of Tgl does not affect its accumulation or assembly. However, like a tgl insertional mutation, the tglC116A allele causes increased extractability of polypeptides of about 40, 28, and 16 kDa in addition to GerQ (20 kDa) and affects the structural integrity of the coat. We show that most Tgl is assembled onto the spore surface soon after its synthesis in the mother cell under sigma(K) control but that the complete insolubilization of at least two of the Tgl-controlled polypeptides occurs several hours later. We also show that a multicopy allele of tgl causes increased assembly of Tgl and affects the assembly, structure, and functional properties of the coat.
authors.affiliation: Instituto de Tecnologia Química e Biológica, Universidade Nova de Lisboa, Oeiras Codex, Portugal.
authors.forename: Rita
authors.initials: R
authors.lastname: Zilhão
authors.forename: Rachele
authors.initials: R
authors.lastname: Isticato
authors.forename: Lígia O
authors.initials: LO
authors.lastname: Martins
authors.forename: Leif
authors.initials: L
authors.lastname: Steil
authors.forename: Uwe
authors.initials: U
authors.lastname: Völker
authors.forename: Ezio
authors.initials: E
authors.lastname: Ricca
authors.forename: Charles P
authors.initials: CP
authors.lastname: Moran
authors.forename: Adriano O
authors.initials: AO
authors.lastname: Henriques
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D001412:Bacillus subtilis / Q000201:enzymology* / Q000648:ultrastructure; D001426:Bacterial Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D017353:Gene Deletion; D018628:Gene Dosage; D015870:Gene Expression; D050939:Gene Fusion; D017930:Genes, Reporter; D046529:Microscopy, Electron, Transmission; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D020125:Mutation, Missense; D017386:Sequence Homology, Amino Acid; D013171:Spores, Bacterial / Q000737:chemistry / Q000201:enzymology* / Q000648:ultrastructure; D011503:Transglutaminases / Q000378:metabolism*; D001616:beta-Galactosidase / Q000032:analysis / Q000235:genetics
chemical_list: D001426:Bacterial Proteins; C077494:CotE protein, Bacillus subtilis; C030069:spore-specific proteins, Bacillus; D011503:Transglutaminases; D001616:beta-Galactosidase