Molecular cloning of the gene encoding beta-1,3(4)-glucanase A from a marine bacterium, Pseudomonas sp. PE2, an essential enzyme for the degradation of Pythium porphyrae cell walls.
The beta-1,3(4)-glucanase A (GluA)-encoding gene named gluA was cloned from the genomic library of a marine bacterium Pseudomonas sp. PE2 by expression in Escherichia coli, and the complete DNA sequence was determined. The recombinant enzyme from Pseudomonas sp. PE2 was examined to determine the essential enzymes for degrading Pythium porphyrae cell walls, comparatively using other two recombinant enzymes, chitinase A and beta-1,3-glucanase B from the same bacterial strain. GluA most degraded the cell walls among these three enzymes, suggesting that GluA seems to be most important to P. porphyrae cell-wall-degrading activity. The deduced GluA is a modular enzyme composed of an N-terminal signal peptide, the tandem-duplicated carbohydrate-binding module family 6 (CBM(GluA)-1 and CBM(GluA)-2), and a glycoside hydrolase family 16 catalytic domain. Deletion analysis clearly indicated that GluA lacking CBM(GluA)-1 and CBM(GluA)-2 does not bind to Avicel and xylan. These results suggest that the tandem-repeated CBM of GluA may play a key role in the binding of Avicel and xylan as well as beta-1,3- and beta-1,3;1,4-glucans and is very important to bind to insoluble polysaccharides.
Kitamura
Etsushi
E
Coastal Bioenvironment Center, Saga University, 152-1 Shonan-cho, Karatsu, Saga 847-0021, Japan.
Kamei
Yuto
Y
D000595:Amino Acid Sequence; D001483:Base Sequence; D002473:Cell Wall / Q000378:metabolism*; D003001:Cloning, Molecular; D006026:Glycoside Hydrolases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D011549:Pseudomonas / Q000201:enzymology* / Q000235:genetics; D011775:Pythium / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D012623:Seawater / Q000382:microbiology*; D017422:Sequence Analysis, DNA; D013379:Substrate Specificity; D047071:beta-Glucans / Q000378:metabolism
D011994:Recombinant Proteins; D047071:beta-Glucans; D006026:Glycoside Hydrolases