The chitin catabolic cascade in the marine bacterium Vibrio furnissii. Molecular cloning, isolation, and characterization of a periplasmic beta-N-acetylglucosaminidase.
We have described some steps in chitin catabolism by Vibrio furnissii, and proposed that chitin oligosaccharides are hydrolyzed in the periplasmic space to GlcNAc and (GlcNAc)2. Since (GlcNAc)2 is an important inducer in the cascade, it must resist hydrolysis in the periplasm. Known V. furnissii periplasmic hydrolases comprise an endoenzyme (Keyhani, N. O. and Roseman, S. (1996) J. Biol. Chem. 271, 33414-33424), and the beta-N-acetylglucosaminidase, ExoI, reported here. ExoI was isolated from a recombinant strain of Escherichia coli, and hydrolyzes aryl-beta-GlcNAc, aryl-beta-GalNAc, and chitin oligosaccharides. No other beta-GlcNAc glycosides were cleaved. The pH optimum was 7.0 for (GlcNAc)n, n = 3-6, but 5.8 for (GlcNAc)2. At the pH of sea water (8.0-8.3), the enzymatic activity with (GlcNAc)2 is virtually undetectable. These results explain the stability of (GlcNAc)2 in the periplasmic space. The cloned beta-GlcNAcidase gene, exoI, encodes a 69,377-kDa protein (611 amino acids); the predicted N-terminal 20 amino acid residues matched those of the isolated protein. The protein amino acid sequence displays significant homologies to the alpha- and beta-chains of human hexosaminidase despite their marked differences in substrate specificities and pH optima.
Keyhani
N O
NO
Department of Biology and the McCollum-Pratt Institute, The Johns Hopkins University, Baltimore, Maryland 21218, USA.
Roseman
S
S
D000118:Acetylglucosaminidase / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D000595:Amino Acid Sequence; D002686:Chitin / Q000378:metabolism*; D003001:Cloning, Molecular; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D009994:Osmolar Concentration; D016415:Sequence Alignment; D013379:Substrate Specificity; D014733:Vibrio / Q000201:enzymology* / Q000378:metabolism
D002686:Chitin; D000118:Acetylglucosaminidase