This work was supported by a Grant-in-aid

for Scientific Research (C) from The Japan Society for the Promotion of Science (No. 19592135 to K.S.). Fig. S1. Verification of inner membrane fractions (IM) and outer membrane ERK inhibitor manufacturer fractions (OM) prepared from W83 (WT), 83K5 (Sov-His), and 83K3 (Δsov). Outer membrane fractions were verified by immnoblot analysis using anti-Pgm6/7 atiserum raised against Pgm6/7 proteins from Porphyromonas gingivalis ATCC33277 [Nagano K, Read EK, Murakami Y, Masuda T, Noguchi T & Yoshimura Y (2005) J Bacteriol187: 902-911]. Inner membrane fractions were verified by checking the NADH-ferricyanide oxidoreductase activity [Futai M (1974) J Membr Biol15: 15-28]. The reduction of ferricyanide was determined in 80 mM Tris-HCl, pH 7.4, 9.0 mM KCN, 1.0 mM NADH, and 0.7 mM ferricyanide, and measured at 420 nm. We thank Fuminobu Yoshimura for the anti-Pgm6/7 antiserum. Please note: Wiley-Blackwell is not responsible for the content or functionality of any supporting materials supplied by the authors. Any queries (other than missing material) should be directed to the corresponding author for the article. “
“It has been demonstrated previously that Enterococcus faecalis produces secreted endoglycosidases that enable the bacteria to remove N-linked glycans from glycoproteins. One enzyme potentially responsible for this activity is EF0114, comprising

a typical GH18 endoglycosidase domain and a GH20 domain. We have analyzed the other candidate, EF2863, selleckchem and show that this predicted single domain GH18 protein is an endo-β-N-acetylglucosaminidase. EF2863 hydrolyzes the glycosidic bond between two N-acetylglucosamines (GlcNAc) in N-linked glycans of the high-mannose and hybrid type, releasing the glycan and leaving one GlcNAc attached to the protein. The activity

of EF2863 is similar to that of the well known deglycosylating enzyme EndoH from Streptomyces plicatus. According to the CAZy nomenclature, the enzyme is designated EfEndo18A. Enterococci are Gram-positive lactic acid bacteria that traditionally have been heptaminol considered harmless inhabitants of the intestinal tract of mammals. However, in recent years, enterococci have emerged as nosocomial pathogens, causing urinary tract infections, bacteraemia and infective endocarditis. Most clinical infections caused by enterococci are due to Enterococcus faecalis (Tendolkar et al., 2003; Fisher & Phillips, 2009). Enterococcus faecalis V583 was the first clinical vancomycin-resistant enterococcal isolate reported in the USA (Sahm et al., 1989) and sequencing of the genome was completed in 2003 (Paulsen et al., 2003). Due to increasing problems with antibiotic resistance, treatment of enterococcal infections is difficult, and there is a need for alternative strategies for overcoming pathogen-related diseases.