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Characterization of β-1,3-galactosyl-N-acetylhexosamine phosphorylase from Propionibacterium acnes

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Homologs of the β-1,3-galactosyl-N-acetylhexosamine phosphorylase (GalHexNAcP) gene (gnpA) were cloned from the genomic DNA of Propionibacterium acnes JCM6425 and P. acnes JCM6473, showing 99.9% and 97.9% nucleotide sequence identity, respectively, with the ppa0083 gene from the genome-sequenced P. acnes KPA171202. No gnpA gene was detected in the genomic DNA of type strain P. acnes ATCC25746. The recombinant enzyme from P. acnes JCM6425 (GnpA) showed approximately 70 times higher specific activity of phosphorolysis on galacto-N-biose (Galβ1→3GalNAc, GNB) than that on lacto-N-biose I (Galβ1→3GlcNAc). K m value for GnpA on GNB was high, but GnpA did not exhibit activity on any derivatives of GNB examined. These results indicate that GnpA is GalHexNAcP which should be classified as galacto-N-biose phosphorylase. The large k cat value of GnpA on GalNAc suggests that GnpA would be a useful catalyst for the synthesis of GNB.

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  1. Allison C, Macfarlane GT (1989) Dissimilatory nitrate reduction by Propionibacterium acnes. Appl Environ Microbiol 55:2899–2903

  2. Bendtsen JD, Nielsen H, von Heijne G, Brunak S (2004) Improved prediction of signal peptides: SignalP 3.0. J Mol Biol 340:783–795

  3. Bruggemann H, Henne A, Hoster F, Liesegang H, Wiezer A, Strittmatter A, Hujer S, Durre P, Gottschalk G (2004) The complete genome sequence of Propionibacterium acnes, a commensal of human skin. Science 305:671–673

  4. Cantarel BL, Coutinho PM, Rancurel C, Bernard T, Lombard V, Henrissat B (2009) Carbohydrate-Active EnZymes database (CAZy): an expert resource for Glycogenomics. Nucleic Acids Res 37:D233–D238

  5. Derensy-Dron D, Krzewinski F, Brassart C, Bouquelet S (1999) β-1,3-Galactosyl-N-acetylhexosamine phosphorylase from Bifidobacterium bifidum DSM 20082: characterization, partial purification and relation to mucin degradation. Biotechnol Appl Biochem 29:3–10

  6. Fujita K, Oura F, Nagamine N, Katayama T, Hiratake J, Sakata K, Kumagai H, Yamamoto K (2005) Identification and molecular cloning of a novel glycoside hydrolase family of core 1 type O-glycan-specific endo-α-N-acetylgalactosaminidase from Bifidobacterium longum. J Biol Chem 280:37415–37422

  7. Henrissat B (1991) A classification of glycosyl hydrolases based on amino-acid sequence similarities. Biochem J 280:309–316

  8. Honda Y, Kitaoka M, Hayashi K (2004) Reaction mechanism of chitobiose phosphorylase from Vibrio proteolyticus: identification of family 36 glycosyltransferase in Vibrio. Biochem J 377:225–232

  9. Huwel S, Haalck L, Conrath N, Spener F (1997) Maltose phosphorylase from Lactobacillus brevis: purification, characterization, and application in a biosensor for ortho-phosphate. Enzyme Microbial Technol 21:413–420

  10. Ishige I, Eishi Y, Takemura T, Kobayashi I, Nakata K, Tanaka I, Nagaoka S, Iwai K, Watanabe K, Takizawa T, Koike M (2005) Propionibacterium acnes is the most common bacterium commensal in peripheral lung tissue and mediastinal lymph nodes from subjects without sarcoidosis. Sarcoidosis Vasc Diffuse Lung Dis 22:33–42

  11. Jakab E, Zbinden R, Gubler J, Ruef C, von Graevenitz A, Krause M (1996) Severe infections caused by Propionibacterium acnes: an underestimated pathogen in late postoperative infections. Yale J Biol Med 69:477–482

  12. Katayama T, Sakuma A, Kimura T, Makimura Y, Hiratake J, Sakata K, Yamanoi T, Kumagai H, Yamamoto K (2004) Molecular cloning and characterization of Bifidobacterium bifidum 1,2-α-l-fucosidase (AfcA), a novel inverting glycosidase (Glycoside hydrolase family 95). J Bacteriol 186:4885–4893

  13. Kitaoka M, Sasaki T, Taniguchi H (1992) Phosphorolytic reaction of Cellvibrio gilvus cellobiose phosphorylase. Biosci Biotechnol Biochem 56:652–655

  14. Kitaoka M, Tian J, Nishimoto M (2005) Novel putative galactose operon involving lacto-N-biose phosphorylase in Bifidobacterium longum. Appl Environ Microbiol 71:3158–3162

  15. Koutsioulis D, Landry D, Guthrie E (2008) Novel endo-α-N-acetylgalactosaminidases with broader substrate specificity. Glycobiology 18:799–805

  16. Kudo T, Iwasaki H, Nishihara S, Shinya N, Ando T, Narimatsu I, Narimatsu H (1996) Molecular genetic analysis of the human Lewis histo-blood group system. J Biol Chem 271:9830–9837

  17. Lowry OH, Lopez JA (1946) The determination of inorganic phosphate in the presence of labile phosphate esters. J Biol Chem 162:421–428

  18. McLorinan GC, Glenn JV, McMullan MG, Patrick S (2005) Propionibacterium acnes wound contamination at the time of spinal surgery. Clin Orthop Relat Res 437:67–73

  19. Mitsuoka T (1996) Intestinal flora and human health. Asia Pac J Clin Nutr 5:2–9

  20. Nagae M, Tsuchiya A, Katayama T, Yamamoto K, Wakatsuki S, Kato R (2007) Structural basis of the catalytic reaction mechanism of novel 1,2-α-l-fucosidase from Bifidobacterium bifidum. J Biol Chem 282:18497–18509

  21. Nakajima M, Kitaoka M (2008) Identification of lacto-N-Biose I phosphorylase from Vibrio vulnificus CMCP6. Appl Environ Microbiol 74:6333–6337

  22. Nakajima M, Nihira T, Nishimoto M, Kitaoka M (2008) Identification of galacto-N-biose phosphorylase from Clostridium perfringens ATCC1312. Appl Microbiol Biotechnol 78:465–471

  23. Nielsen H, Engelbrecht J, Brunak S, von Heijne G (1997) Identification of prokaryotic and eukaryotic signal peptides and prediction of their cleavage sites. Protein Eng 10:1–6

  24. Nihira T, Nakajima M, Inoue K, Nishimoto M, Kitaoka M (2007) Colorimetric quantification of α-d-galactose 1-phosphate. Anal Biochem 371:259–261

  25. Nishimoto M, Kitaoka M (2007a) Identification of the putative proton donor residue of lacto-N-biose phosphorylase (EC Biosci Biotechnol Biochem 71:1587–1591

  26. Nishimoto M, Kitaoka M (2007b) Practical preparation of Lacto-N-biose I, the candidate of the bifidus factor in human milk. Biosci Biotechnol Biochem 71:2101–2104

  27. Nishimoto M, Kitaoka M (2007c) The complete lacto-N-biose I/galacto-N-biose metabolic pathway in Bifidobacterium longum: identification of N-acetylhexosamine 1-kinase. Appl Environ Microbiol 73:6444–6449

  28. Suzuki R, Wada J, Katayama T, Fushinobu S, Wakagi T, Shoun H, Sugimoto H, Tanaka A, Kumagai H, Ashida H, Kitaoka M, Yamamoto K (2008) Structural and thermodynamic analyses of solute-binding protein from Bifidobacterium longum specific for core 1 disaccharide and lacto-N-biose I. J Biol Chem 283:13165–13173

  29. Wada J, Suzuki R, Fushinobu S, Kitaoka M, Wakagi T, Shoun H, Ashida H, Kumagai H, Katayama T, Yamamoto K (2007) Purification, crystallization and preliminary X-ray analysis of the galacto-N-biose-/lacto-N-biose I-binding protein (GL-BP) of the ABC transporter from Bifidobacterium longum JCM1217. Acta Cryst F 63:751–753

  30. Wada J, Ando T, Kiyohara M, Ashida H, Kitaoka M, Yamaguchi M, Kumagai H, Katayama T, Yamamoto K (2008) Bifidobacterium bifidum lacto-N-biosidase, a critical enzyme for the degradation of human milk oligosaccharides with a type 1 structure. Appl Environ Microbiol 74:3996–4004

  31. Yamada T, Eishi Y, Ikeda S, Ishige I, Suzuki T, Takemura T, Takizawa T, Koike M (2002) In situ localization of Propionibacterium acnes DNA in lymph nodes from sarcoidosis patients by signal amplification with catalysed reporter deposition. J Pathol 198:541–547

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Correspondence to Motomitsu Kitaoka.

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Nakajima, M., Nishimoto, M. & Kitaoka, M. Characterization of β-1,3-galactosyl-N-acetylhexosamine phosphorylase from Propionibacterium acnes . Appl Microbiol Biotechnol 83, 109–115 (2009).

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  • EC
  • Galacto-N-biose phosphorylase
  • Galacto-N-biose
  • Propionibacterium acnes
  • Mucin
  • Lacto-N-biose I