Department of Chemical Engineering, University of Texas at Austin, Austin, Texas, USA.
Nat Chem Biol. 2011 Feb;7(2):101-5. doi: 10.1038/nchembio.499. Epub 2010 Dec 26.
The first and highly conserved step in glutathione (GSH) biosynthesis is formation of γ-glutamyl cysteine by the enzyme glutamate-cysteine ligase (GshA). However, bioinformatic analysis revealed that many prokaryotic species that encode GSH-dependent proteins lack the gene for this enzyme. To understand how bacteria cope without gshA, we isolated Escherichia coli ΔgshA multigenic suppressors that accumulated physiological levels of GSH. Mutations in both proB and proA, the first two genes in L-proline biosynthesis, provided a new pathway for γ-glutamyl cysteine formation via the selective interception of ProB-bound γ-glutamyl phosphate by amino acid thiols, likely through an S-to-N acyl shift mechanism. Bioinformatic analysis suggested that the L-proline biosynthetic pathway may have a second role in γ-glutamyl cysteine formation in prokaryotes. Also, we showed that this mechanism could be exploited to generate cytoplasmic redox buffers bioorthogonal to GSH.
谷胱甘肽（GSH）生物合成的第一步也是高度保守的一步是由谷氨酸-半胱氨酸连接酶（GshA）形成γ-谷氨酰半胱氨酸。然而，生物信息学分析表明，许多编码 GSH 依赖性蛋白的原核生物物种缺乏该酶的基因。为了了解没有 gshA 的细菌如何应对，我们分离出了大肠杆菌 ΔgshA 多基因抑制剂，该抑制剂积累了生理水平的 GSH。脯氨酸生物合成的前两个基因 proB 和 proA 中的突变为γ-谷氨酰半胱氨酸的形成提供了一条新途径，通过选择性截获 ProB 结合的γ-谷氨酰磷酸，可能通过 S 到 N 酰基转移机制。生物信息学分析表明，L-脯氨酸生物合成途径在原核生物中可能具有形成γ-谷氨酰半胱氨酸的第二个作用。此外，我们还表明，该机制可用于产生与 GSH 生物正交的细胞质氧化还原缓冲剂。
