Characterization of a novel glucosamine-6-phosphate deaminase from a hyperthermophilic archaeon
Characterization of a novel glucosamine-6-phosphate deaminase from a hyperthermophilic archaeon
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DOI：
10.1128/jb.187.20.7038-7044.2005
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发表时间：
2005-10-01
影响因子：
3.2
通讯作者：
Imanaka, T
中科院分区：
文献类型：
--
作者：
Tanaka, T;Takahashi, F;Imanaka, T
氨基糖代谢的关键步骤是果糖-6-磷酸（Fru6P）和氨基葡萄糖-6-磷酸（GIcN6P）之间的相互转化。这种转化分别由GlcN6P脱氨酶和GlcN6P合成酶在分解代谢和合成代谢方向上催化，这两种酶在初级结构上没有相互关系。在这项研究中，我们研究了超嗜热古细菌柯达卡拉热球菌（Thermococcus kodakaraensis） KOD1中存在于几丁质降解基因簇中的glmD基因产物（glmD (Tk)）的催化特性和调控特征。虽然GlmD（Tk）蛋白被预测可能是与GlcN6P合成酶c端糖异构酶结构域相关的糖异构酶，但重组GlmD（Tk）明确表现出GlcN6P脱氨酶活性，从GlcN6P产生Fru6P和氨。该酶还催化了逆反应，即氨依赖的Fru6P胺化/异构化为GIcN6P，而没有观察到GlcN6P合成酶依赖谷氨酰胺的活性。动力学分析表明，该酶更倾向于脱氨酶反应，而不是逆向反应，这与GlmD（Tk）的分解代谢功能一致。在T. kodakaraensis细胞中，glmD（Tk）与上游编码ABC转运蛋白的基因和下游编码β -氨基葡萄糖苷酶外显子基因（glmA(Tk)）一起多顺反转录，并由几丁质降解中间体二乙酰糖诱导表达。研究结果表明，GlmD（Tk）实际上是一种GlcN6P脱氨酶，在这种嗜热细菌中，几丁质衍生的单糖进入糖酵解过程中起作用。该酶是古细菌GIcN6P脱氨酶的第一个例子，是一种结构新颖的类型，不同于任何已知的GlcN6P脱氨酶。
A key step in amino sugar metabolism is the interconversion between fructose-6-phosphate (Fru6P) and glucosamine-6-phosphate (GIcN6P). This conversion is catalyzed in the catabolic and anabolic directions by GlcN6P deaminase and GlcN6P synthase, respectively, two enzymes that show no relationship with one another in terms of primary structure. In this study, we examined the catalytic properties and regulatory features of the glmD gene product (GlmD(Tk)) present within a chitin degradation gene cluster in the hyperthermophilic archaeon Thermococcus kodakaraensis KOD1. Although the protein GlmD(Tk) was predicted as a probable sugar isomerase related to the C-terminal sugar isomerase domain of GlcN6P synthase, the recombinant GlmD(Tk), clearly exhibited GlcN6P deaminase activity, generating Fru6P and ammonia from GlcN6P. This enzyme also catalyzed the reverse reaction, the ammonia-dependent amination/isomerization of Fru6P to GIcN6P, whereas no GlcN6P synthase activity dependent on glutamine was observed. Kinetic analyses clarified the preference of this enzyme for the deaminase reaction rather than the reverse one, consistent with the catabolic function of GlmD(Tk). In T. kodakaraensis cells, glmD(Tk) was polycistronically transcribed together with upstream genes encoding an ABC transporter and a downstream exo-beta-glucosaminidase gene (glmA(Tk)) within the gene cluster, and their expression was induced by the chitin degradation intermediate, diacetylehitobiose. The results presented here indicate that GlmD(Tk) is actually a GlcN6P deaminase functioning in the entry of chitin-derived monosaccharides to glycolysis in this hyperthermophile. This enzyme is the first example of an archaeal GIcN6P deaminase and is a structurally novel type distinct from any previously known GlcN6P deaminase.