Molecular Cloning, Sequencing, and Expression of a l-Glutamine d-Fructose 6-Phosphate Amidotransferase Gene from Volvariella volvacea
文献类型: 外文期刊
第一作者: Luo, Chuping
作者: Luo, Chuping;Chen, Zhiyi;Liu, Yongfeng;Luo, Chuping;Shao, Weilan;Li, Xun;Li, Xun
作者机构:
关键词: L-Glutamine:D-fructose-6-phosphate amidotransferase;Volvariella volvacea;cDNA cloning;Heterologous expression
期刊名称:PROTEIN JOURNAL ( 影响因子:2.371; 五年影响因子:1.929 )
ISSN: 1572-3887
年卷期: 2009 年 28 卷 1 期
页码:
收录情况: SCI
摘要: Using 3'-RACE and 5'-RACE, we have cloned and sequenced the genomic gene and complete cDNA encoding l-glutamine d-fructose 6-phosphate amidotransferase (GFAT) from the edible straw mushroom, Volvariella volvacea. Gfat contains five introns, and encodes a predicted protein of 697 amino acids that is homologous to other reported GFAT sequences. Southern hybridization indicated that a single gfat gene locus exists in the V. volvacea genome. Recombinant native V. volvacea GFAT enzyme, over-expressed using Escherichia coli and partially purified, had an estimated molecular mass of 306 kDa and consisted of four equal-sized subunits of 77 kD. Reciprocal plots revealed K (m) values of 0.55 and 0.75 mM for fructose 6-phosphate and l-glutamine, respectively. V. volvacea GFAT activity was inhibited by the end-product of the hexosamine pathway, UDP-GlcNAc, and by the glutamine analogues N (3)-(4-methoxyfumaroyl)-l-2,3-diaminopropanoic acid and 2-amino-2-deoxy-d-glucitol-6-phosphate.
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