A 383Thr→Ala Mutation Elevates the β-galactosidase Activity Resulting in a Lac+ Phenotype in Xanthom

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Title:	A 383Thr→Ala Mutation Elevates the β-galactosidase Activity Resulting in a Lac+ Phenotype in Xanthom
Authors:	楊翠青(Yang,Tsuey-Ching)
Contributors:	健康照護學院醫學檢驗生物技術學系
Date:	2001-08-21
Issue Date:	2009-09-04 15:49:36 (UTC+8)
Abstract:	The species Xanthomonas campestris, which causes systemic black vein and rot of crucifers, comprises over 125 different pathovars. In addition to phenotypic, chemical and genetic approaches, the taxonomy of genus Xanthomonas has been suggested by the metabolic activity of the bacteria on 95 different carbon compounds, including lactose. Although b-galactosidase, the enzyme required for lactose catabolism, has been isolated from X. campestris pv. campestris, this bacterium scarcely grows on lactose medium owing to low levels of the enzyme. Therefore, this pathovars has been classified as Lac-. In this study, a lactose-utilizing Xc17 mutant strain, designed as Xc17L, was isolated after NaNO2 mutagenesis. With a b-galactosidase level 3.5-fold higher than that in Xc17, Xc17L grew in the lactose medium at a similar rate to that with glucose. The DNA fragment containing the Xc17 b-galactosidase gene, encoding 613 amino acid residues with a calculated MW of 68 kDa, was cloned and sequenced. Using a PCR-based strategy, the mutant gene was also cloned from Xc17L and sequenced. Sequence comparison revealed that three nucleotides were altered, of which only one resulted in a 383Thr?Ala mutation. No mutation was found in the 800-bp upstream region presumably containing the promoter sequences. Xc17 containing the cloned b-galactosidase gene expressed an enzyme level similar to that of Xc17L and was able to grow in lactose medium, indicating that elevation of the b-galactosidase activity in Xc17L is associated with the 383Thr?Ala alteration. The results of this study have thus changed the physiological characteristics of X. campestris pv. campestris from Lac- to Lac+.
Relation:	The 9th SCBA International Symposium.
Appears in Collections:	[Graduate Institute of Nutrition ] Proceedings

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