Single-step pathway for synthesis of glucosylglycerate in Persephonella marina
Single-step pathway for synthesis of glucosylglycerate in Persephonella marina
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DOI：
10.1128/jb.00075-07
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发表时间：
2007-06-01
影响因子：
3.2
通讯作者：
da Costa, Milton S.
中科院分区：
文献类型：
--
作者：
Fernandes, Chantal;Empadinhas, Nuno;da Costa, Milton S.
基于海洋大隐藻重组葡萄糖甘油酸合酶（Ggs）的活性，提出了一条一步合成相容性溶质葡萄糖甘油酸（GG）的途径。相应的基因编码一种推定的糖基转移酶，它是操纵子样结构的一部分，该结构还包含葡糖基-3-磷酸甘油酸合酶（GpgS）和葡糖基-3-磷酸甘油酸磷酸酶（GpgP）的基因，这些酶通过形成葡糖基-3-磷酸甘油酸导致GG的合成。推定的葡糖基转移酶基因在大肠杆菌中表达，并且重组产物催化从ADP-葡萄糖和D-甘油酸一步合成GG，在70 ° C下的Km值分别为1.5和2.2 mM。该葡糖基甘油酸合酶（Ggs）也能够使用GDP-和UDP-葡萄糖作为供体来形成GG，但效率较低。在80至85 ℃的温度下观察到最大活性，并且催化需要Mg 2+或Ca 2+。Ggs活性最大，在pH值5.5和pH 8.0之间几乎保持恒定，85 ℃下的灭活半衰期为74 h，100 ℃下为8 min。这是第一次报告的酶催化合成的GG在一个步骤和存在两个途径的GG合成在同一个有机体。
A single-step pathway for the synthesis of the compatible solute glucosylglycerate (GG) is proposed based on the activity of a recombinant glucosylglycerate synthase (Ggs) from Persephonella marina. The corresponding gene encoded a putative glycosyltransferase that was part of an operon-like structure which also contained the genes for glucosyl-3-phosphoglycerate synthase (GpgS) and glucosyl-3-phosphoglycerate phosphatase (GpgP), the enzymes that lead to the synthesis of GG through the formation of glucosyl-3-phosphoglycerate. The putative glucosyltransferase gene was expressed in Escherichia coli, and the recombinant product catalyzed the synthesis of GG in one step from ADP-glucose and D-glycerate, with K-m values at 70 degrees C of 1.5 and 2.2 mM, respectively. This glucosylglycerate synthase (Ggs) was also able to use GDP- and UDP-glucose as donors to form GG, but the efficiencies were lower. Maximal activity was observed at temperatures between 80 and 85 degrees C, and Mg2+ or Ca2+ was required for catalysis. Ggs activity was maximal and remained nearly constant at pH values between 5.5 and pH 8.0, and the half-lives for inactivation were 74 h at 85 degrees C and 8 min at 100 degrees C. This is the first report of an enzyme catalyzing the synthesis of GG in one step and of the existence of two pathways for GG synthesis in the same organism.