Chitin catabolism in the marine bacterium Vibrio furnissii. Identification and molecular cloning of a chitoporin.
Chitin catabolism by the marine bacterium Vibrio furnissii involves many genes and proteins, including two unique periplasmic hydrolases, a chitodextrinase and a beta-N-acetylglucosaminidase (Keyhani, N. O. , and Roseman, S. (1996) J. Biol. Chem. 271, 33414-33424 and 33425-33432). A specific chitoporin in the outer membrane may be required for these glycosidases to be accessible to extracellular chitooligosaccharides, (GlcNAc)(n), that are produced by chitinases. We report here the identification and molecular cloning of such a porin. An outer membrane protein, OMP (apparent molecular mass 40 kDa) was expressed when V. furnissii was induced by (GlcNAc)(n), n = 2-6, but not by GlcNAc or other sugars. Based on the N-terminal sequence of OMP, oligonucleotides were synthesized and used to clone the gene, chiP. The deduced amino acid sequence of ChiP is similar to several bacterial porins; OMP is a processed form of ChiP. In Escherichia coli, two recombinant proteins were observed, corresponding to processed and unprocessed forms of ChiP. A null mutant of chiP was constructed in V. furnissii. In contrast to the parental strain, the mutant did not grow on (GlcNAc)(3) and transported a nonmetabolizable analogue of (GlcNAc)(2) at a reduced rate. These results imply that ChiP is a specific chitoporin.
Keyhani
N O
NO
Department of Biology and the McCollum-Pratt Institute, The Johns Hopkins University, Baltimore, Maryland 21218, USA.
Li
X B
XB
Roseman
S
S
D000595:Amino Acid Sequence; D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D001483:Base Sequence; D050260:Carbohydrate Metabolism; D002462:Cell Membrane / Q000378:metabolism; D002686:Chitin / Q000378:metabolism*; D003001:Cloning, Molecular; D003227:Conjugation, Genetic; D004926:Escherichia coli / Q000235:genetics; D015964:Gene Expression Regulation, Bacterial; D005816:Genetic Complementation Test; D007700:Kinetics; D008969:Molecular Sequence Data; D018272:Porins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D012623:Seawater / Q000382:microbiology; D014733:Vibrio / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*
D001425:Bacterial Outer Membrane Proteins; D018272:Porins; D011994:Recombinant Proteins; C415833:chiP protein, Vibrio furnissii; D002686:Chitin