HemK, a class of protein methyl transferase with similarity to DNA methyl transferases, methylates polypeptide chain release factors, and hemK knockout induces defects in translational termination
HemK, a class of protein methyl transferase with similarity to DNA methyl transferases, methylates polypeptide chain release factors, and hemK knockout induces defects in translational termination
复制标题
DOI：
10.1073/pnas.032488499
复制
发表时间：
2002-02-05
影响因子：
11.1
通讯作者：
Inokuchi, H
中科院分区：
文献类型：
--
作者：
Nakahigashi, K;Kubo, N;Inokuchi, H
HemK是一种功能未知的普遍保守蛋白，与DNA-（腺嘌呤- n6）甲基转移酶（MTases）具有高度的氨基酸相似性。hemK基因的某种突变挽救了大肠杆菌中铁螯合酶缺陷（hemH）突变体的光敏表型。通过微阵列分析发现，hemK敲除菌株大肠杆菌不仅存在严重的生长缺陷，而且基因表达全局向厌氧呼吸转变，这种转变可能通过减少氧化应激而导致光敏性的丧失。抑制突变消除了hemK敲除菌株的生长缺陷，并被证明是由多肽链释放因子（RF） 2的密码子246上的苏氨酸到丙氨酸的变化引起的，表明hemK在翻译终止中起作用。与这种作用相一致的是，hemK敲除菌株显示出无义密码子的穿透率提高，并诱导细胞内蛋白质的转移mrna介导的标记。通过对体内和体外RF1和RF2的甲基化分析，我们发现HemK在体外含有保守GGQ基序的胰蛋白酶片段中甲基化RF1和RF2，并且HemK至少是体内RF1相同片段中甲基化所必需的。这是一个含有DNA MTase基序的蛋白质AATase和蛋白质-（谷氨酰胺- n5） MTase的例子。
HemK, a universally conserved protein of unknown function, has high amino acid similarity with DNA-(adenine-N6) methyl transferases (MTases). A certain mutation in hemK gene rescues the photosensitive phenotype of a ferrochelatase-deficient (hemH) mutant in Escherichia coli. A hemK knockout strain of E coli not only suffered severe growth defects, but also showed a global shift in gene expression to anaerobic respiration, as determined by microarray analysis, and this shift may lead to the abrogation of photosensitivity by reducing the oxidative stress. Suppressor mutations that abrogated the growth defects of the hemK knockout strain were isolated and shown to be caused by a threonine to alanine change at codon 246 of polypeptide chain release factor (RF) 2, indicating that hemK plays a role in translational termination. Consistent with such a role, the hemK knockout strain showed an enhanced rate of read-through of nonsense codons and induction of transfer-mRNA-mediated tagging of proteins within the cell. By analysis of the methylation of RF1 and RF2 in vivo and in vitro, we showed that HemK methylates RF1 and RF2 in vitro within the tryptic fragment containing the conserved GGQ motif, and that hemK is required for the methylation within the same fragment of, at least, RF1 in vivo, This is an example of a protein AATase containing the DNA MTase motif and also a protein-(glutamine-N5) MTase.