A novel acetyltransferase found in Saccharomyces cerevisiae Σ1278b that detoxifies a proline analogue, azetidine-2-carboxylic acid
A novel acetyltransferase found in Saccharomyces cerevisiae Σ1278b that detoxifies a proline analogue, azetidine-2-carboxylic acid
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DOI：
10.1074/jbc.c100487200
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发表时间：
2001-11-09
影响因子：
4.8
通讯作者：
Takagi, H
中科院分区：
文献类型：
--
作者：
Shichiri, M;Hoshikawa, C;Takagi, H
L-氮杂环丁烷-2-羧酸（Azetidine-2-carboxylic acid，AZC）是L-脯氨酸的一种毒性四元环类似物，通过脯氨酸转运蛋白转运到细胞内。它引起蛋白质的错误折叠，它与L-脯氨酸竞争性地掺入蛋白质中，从而抑制细胞的生长。我们最近在酿酒酵母Sigma 1278 b的染色体上发现了一个新的基因MPR 1，该基因是Sigma 1278背景菌株对有毒AZC的抗性所必需的。该基因在用于基因组序列测定的特定酵母菌株中缺失。虽然该蛋白质序列与S.酿酒酵母转录调节因子Mpr 1 p不影响参与脯氨酸摄取的基因的表达。然而，在大肠杆菌中的基因表达和酶分析表明，MPR 1基因编码一种新的AZC乙酰转移酶，通过该酶，L-脯氨酸本身和其他L-脯氨酸类似物不被乙酰化。Mpr 1 p被认为是N-乙酰基转移酶超家族的成员，这是基于通过超家族成员中参与结合乙酰辅酶A的高度保守区域进行Ala扫描诱变的结果。我们的研究结果表明，Mpr 1 p解毒AZC乙酰化它在细胞质中。这种酶可用作多种生物体中的选择性标记，因为表达MPR 1基因的细胞获得AZC抗性表型。
L-Azetidine-2-carboxylic acid (AZC), a toxic four-membered ring analogue Of L-proline, is transported into the cells via proline transporters. It causes misfolding of the proteins into which it is incorporated competitively with L-proline and thereby inhibits the growth of the cells. We recently have discovered, on the chromosome of Saccharomyces cerevisiae Sigma 1278b, a novel gene MPR1 required for the resistance of Sigma 1278 background strains to toxic AZC. This gene was missing in the particular yeast strain used for the genomic sequence determination. Although the protein sequence was homologous to that of the S. cerevisiae transcriptional regulator, Mpr1p did not affect the expression of genes involved in proline uptake. However, gene expression in Escherichia coli and enzymatic analysis showed that the MPR1 gene encodes a novel AZC acetyltransferase, by which L-proline itself and other L-proline analogues are not acetylated. Mpr1p was considered to be a member of the N-acetyl-transferase superfamily based on the results of an Ala-scan mutagenesis through the highly conserved region involved in binding acetyl-CoA in members of the superfamily. Our findings suggest that Mpr1p detoxifies AZC by acetylating it in the cytoplasm. This enzyme might be utilized as a selective marker in a wide variety of organisms, because the cells expressing the MPR1 gene acquire the AZC-resistant phenotype.