Amino-terminal protein processing in Saccharomyces cerevisiae is an essential function that requires two distinct methionine aminopeptidases
Amino-terminal protein processing in Saccharomyces cerevisiae is an essential function that requires two distinct methionine aminopeptidases
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DOI：
10.1073/pnas.92.26.12357
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发表时间：
1995-12-19
影响因子：
11.1
通讯作者：
Chang, YH
中科院分区：
文献类型：
--
作者：
Li, X;Chang, YH
我们之前描述了酿酒酵母中的甲硫氨酸氨肽酶（EC 3.4.11.18；Met-AP1；也称为肽酶 M），其与其原核同源物的不同之处在于，它（i）包含 N 端锌指结构域，并且（ii）在被破坏时不会产生致死性，尽管它确实会显着减慢生长；它由 MAP1 基因编码。在这里，我们描述了酿酒酵母中的第二种甲硫氨酸氨肽酶（Met-AP2），由 MAP2 编码，它被克隆为 map1 缺失菌株缓慢生长表型的抑制因子。 MAP2 的 DNA 序列编码 421 个氨基酸的蛋白质，与酵母 Met-AP1 的序列有 22% 的同一性。令人惊讶的是，与GenBank数据库中的序列比较表明，MAP2的产物与大鼠p(67)具有更高的同源性(55%同一性)，大鼠p(67)被表征为起始因子2相关蛋白，但尚未显示具有Met-AP活性。在高拷贝数质粒中表达 MAP2 的 map1 null 细胞的转化体在不同肽底物上的 Met-AP 活性增加了 3 至 12 倍。通过免疫亲和层析纯化带有表位标记的抑制基因产物，并显示其含有 Met-AP 活性；为了评估 Met-AP2 的生理意义，从野生型和 map1 无效酵母菌株中删除了 MAP2 基因。与map1无效菌株一样，map2无效菌株是可行的，但生长速度较慢。 map1、map2 双无效菌株无法存活。因此，与原核生物一样，N-末端甲硫氨酸的去除是酵母中的基本功能，但酵母需要两种甲硫氨酸氨肽酶来提供仅可由Met-AP1或Met-AP2单独提供的部分功能。
We previously characterized a methionine aminopeptidase (EC 3.4.11.18; Met-AP1; also called peptidase M) in Saccharomyces cerevisiae, which differs from its prokaryotic homologues in that it (i) contains an N-terminal zinc-finger domain and (ii) does not produce lethality when disrupted, although it does slow growth dramatically; it is encoded by a gene called MAP1. Here we describe a second methionine aminopeptidase (Met-AP2) in S. cerevisiae, encoded by MAP2, which was cloned as a suppressor of the slow-growth phenotype of the map1 null strain. The DNA sequence of MAP2 encodes a protein of 421 amino acids that shows 22% identity with the sequence of yeast Met-AP1. Surprisingly, comparison with sequences in the GenBank data base showed that the product of MAP2 has even greater homology (55% identity) with rat p(67), which was characterized as an initiation factor 2-associated protein but not yet shown to have Met-AP activity. Transformants of map1 null cells expressing MAP2 in a high-copy-number plasmid contained 3-to 12-fold increases in Met-AP activity on different peptide substrates. The epitope-tagged suppressor gene product was purified by immunoaffinity chromatography and shown to contain Met-AP activity; To evaluate the physiological significance of Met-AP2, the MAP2 gene was deleted from wild-type and map1 null yeast strains. The map2 null strain, like the map1 null strain, is viable but with a slower growth rate. The map1, map2 double-null strains are nonviable. Thus, removal of N-terminal methionine is an essential function in yeast, as in prokaryotes, but yeast require two methionine aminopeptidases to provide the essential function which can only be partially provided by Met-AP1 or Met-AP2 alone.