野油菜黄单胞菌中长链3-酮脂酰 ACP 合成酶的鉴定
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国家自然科学基金(31200028);高等学校博士学科点专项科研基金(20104404110005);广东省高校优秀青年创新人才培养计划(LYM10038)


Characterization of long chain 3-ketoacyl-ACP synthase in Xanthomonas campestris
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    摘要:

    【目的】研究野油菜黄单胞菌 Xanthomonas campestris 8004基因组中3个标注为3-酮脂酰ACP合成酶的基因XcfabF1XcfabF2XcfabB 在脂肪酸合成过程中的功能.【方法】将这3个基因分别克隆到表达载体pBAD24M,然后转化大肠埃希菌 Escherichia colifabBfabF 温敏型突变株CY242和CY244,同时利用体外无细胞抽提物酶学分析FabF1、FabF2和FabB蛋白活性.【结果和结论】XcfabF1XcfabB 基因分别能遗传互补大肠埃希菌 fabFfabB 突变,而 XcfabF2 基因则不能互补大肠埃希菌 fabFfabB 突变.体外无细胞抽提物酶学分析表明FabF1和FabB都能完成辛脂酰-ACP的延伸,但FabF2则不能完成该酶促反应.XcfabB 基因编码3-酮脂酰ACP合成酶I,XcfabF1 基因编码3-酮脂酰ACP合成酶II,而fabF2 基因不参与中长链脂肪酸的合成.

    Abstract:

    【Objective】The functions of the three genes XcfabF1,XcfabF2 and XcfabB, which were annotated as encodes putative 3-ketoacyl-ACP synthase in fatty acid synthesis in Xanthomonas campestris 8004 genome, were studied. 【Method】Three genes coloned into expression plasmid pBAD24M separately were transformed into the fabB and fabF temperature sensitive mutant CY242 and CY244 of Escherichia coli. The activity of XcFabF1, XcFabF2 and XcFabB in vitro was assayed by cell-free extracts. 【Result and conclusion】The genetic complementary revealed that XcfabB and XcfabF1 genes could restore the growth of mutant CY242 and CY244 respectively, but XcfabF2 could not restore the growth of mutant CY242 or CY244. The cell-free extracts of XcFabF1 and XcFabB could elongate octanoyl-ACP to longer acyl-ACP, while XcFabF2 could not catalyse the elongation in vitro assay. The above results demonstrate that XcfabB and XcfabF1 encode 3-ketoacyl-ACP sysnthase I and 3-ketoacyl-ACP sysnthase II respectively, but XcfabF2 is not involved in long chain fatty acid synthesis in X. Campestris.

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董会娟,余永红,王海洪,马金成.野油菜黄单胞菌中长链3-酮脂酰 ACP 合成酶的鉴定[J].华南农业大学学报,2015,36(2):49-54

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  • 收稿日期:2014-01-01
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  • 在线发布日期: 2015-02-13