Applied Microbiology and Biotechnology

, Volume 60, Issue 4, pp 420–427 | Cite as

A novel β-N-acetylglucosaminidase of Clostridiumparaputrificum M-21 with high activity on chitobiose

  •  H. Li
  •  K. Morimoto
  •  N. Katagiri
  •  T. Kimura
  •  K. Sakka
  •  S. Lun
  •  K. Ohmiya
Original Paper

Abstract.

A β-N-acetylglucosaminidase gene (nag3A) from Clostridium paraputrificum M-21 was cloned in Escherichia coli. The nag3A gene consists of an open reading frame of 1,239-bp, encoding 413 amino acids with a deduced molecular weight of 45,531 Da. Nag3A is a single domain enzyme containing a family 3 glycoside hydrolase catalytic domain. Nag3A was purified from recombinant E. coli and characterized. The enzyme hydrolyzed chitooligomers such as di-N-acetylchitobiose, tri-N-acetylchitotriose, tetra-N-acetylchitotetraose, penta-N-acetylchitopentaose, hexa-N-acetylchitohexaose, ball-milled chitin, and synthetic substrates such as 4-methylumbelliferyl N-acetyl β-D-glucosaminide [4-MU-(GlcNAc)], but had no activity at all against p-nitrophenyl-β-D-glucoside, p-nitrophenyl-β-D-xyloside, or p-nitrophenyl-β-D-galactosamine. The enzyme was optimally active at 50°C and pH 7.0, and the apparent Km and Vmax values for 4-MU-(GlcNAc) were 7.9 µM and 21.8 µmol min–1 mg protein–1, respectively. SDS-PAGE, zymogram, and immunological analyses suggested that this enzyme is induced by ball-milled chitin.

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

Copyright information

© Springer-Verlag 2002

Authors and Affiliations

  •  H. Li
    • 1
  •  K. Morimoto
    • 2
  •  N. Katagiri
    • 3
  •  T. Kimura
    • 2
  •  K. Sakka
    • 2
  •  S. Lun
    • 1
  •  K. Ohmiya
    • 2
  1. 1.School of Biotechnology, Southern Yangtze University, 170 Huihe Road, Wuxi 214036, China
  2. 2.Faculty of Bioresources, Mie University, Kamihama 1515, Tsu 514–8507, Japan
  3. 3.Department of Chemical Engineering, Nagoya University, Furo-cho, Chikusa-ku, Nagoya-City, 464–8603, Japan

Personalised recommendations