title: Characterization of two new glycosyl hydrolases from the lactic acid bacterium Carnobacterium piscicola strain BA.
abstract: Three genes with homology to glycosyl hydrolases were detected on a DNA fragment cloned from a psychrophilic lactic acid bacterium isolate, Carnobacterium piscicola strain BA. A 2.2-kb region corresponding to an alpha-galactosidase gene, agaA, was followed by two genes in the same orientation, bgaB, encoding a 2-kb beta-galactosidase, and bgaC, encoding a structurally distinct 1.76-kb beta-galactosidase. This gene arrangement had not been observed in other lactic acid bacteria, including Lactococcus lactis, for which the genome sequence is known. To determine if these sequences encoded enzymes with alpha- and beta-galactosidase activities, we subcloned the genes and examined the enzyme properties. The alpha-galactosidase, AgaA, hydrolyzes para-nitrophenyl-alpha-D-galactopyranoside and has optimal activity at 32 to 37 degrees C. The beta-galactosidase, BgaC, has an optimal activity at 40 degrees C and a half-life of 15 min at 45 degrees C. The regulation of these enzymes was tested in C. piscicola strain BA and activity on both alpha- and beta-galactoside substrates decreased for cells grown with added glucose or lactose. Instead, an increase in activity on a phosphorylated beta-galactoside substrate was found for the cells supplemented with lactose, suggesting that a phospho-galactosidase functions during lactose utilization. Thus, the two beta-galactosidases may act synergistically with the alpha-galactosidase to degrade other polysaccharides available in the environment.
authors.affiliation: Department of Biochemistry and Microbiology, Rutgers University, New Brunswick, New Jersey 08901-8525, USA.
authors.forename: J
authors.initials: J
authors.lastname: Coombs
authors.forename: J E
authors.initials: JE
authors.lastname: Brenchley
mesh_terms: D001426:Bacterial Proteins; D003001:Cloning, Molecular; D003470:Culture Media; D015964:Gene Expression Regulation, Bacterial; D007777:Lactobacillaceae / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development; D008969:Molecular Sequence Data; D017422:Sequence Analysis, DNA; D013696:Temperature; D000519:alpha-Galactosidase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D001616:beta-Galactosidase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D003470:Culture Media; C444001:BgaC protein, Carnobacterium piscicola; C444000:AgaA protein, Carnobacterium piscicola; D000519:alpha-Galactosidase; D001616:beta-Galactosidase