Xc17L, a lactose-utilizing mutant of Xanthomonas campestris pv. campestris previously isolated by mutagenesis with nitrous acid, displays a level of β-galactosidase 3.5-fold higher than that in the parental Xc17. In this study, the gene encoding the enzyme displaying a higher specific activity in Xc17L was inactivated by mini-Tn5 transposition.Sequencing revealed that the product (579 aa, 63.5 kDa) of this gene, designated galD, was previously annotated to encode a hypothetical protein on the genome. Mutation of the gene by marker exchange, complementation test and Western blot analysis together confirmed that galD is indeed the gene involved in β-galactosidase elevation in Xc17L. With only the N-terminal region possessing similarity to the known β-galactosidases and partially conserved consensus motif, GalD is recognized as a member of the glycosyl hydrolase family 35. Insertion with GmΩ, which causes polar effects, into the upstream genes followed by Western blotting showed that galD is cotranscribed with the upstream genes and expressed constitutively. Mutation in galD causes no significant changes including pathogenicity in the bacterium.
Relation:
The Journal of Molecular Microbiology and Biotechnology 13:172-180
