Cfr and RlmN contain a single [4Fe-4S] cluster, which directs two distinct reactivities for S-adenosylmethionine: methyl transfer by SN2 displacement and radical generation.
Cfr and RlmN contain a single [4Fe-4S] cluster, which directs two distinct reactivities for S-adenosylmethionine: methyl transfer by SN2 displacement and radical generation.
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DOI：
10.1021/ja207327v
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发表时间：
2011-12-14
影响因子：
15
通讯作者：
Booker, Squire J.
中科院分区：
文献类型：
--
作者：
Grove, Tyler L.;Radle, Matthew I.;Krebs, Carsten;Booker, Squire J.
自由基SAM（RS）蛋白RlmN和Cfr分别催化23 S rRNA中腺苷2503的碳2和8的甲基化。这两种反应的范围是相似的，需要合成的甲基部分来源于S-腺苷甲硫氨酸（SAM）亲电子sp2杂化碳原子上通过中间的蛋白质S-甲基半胱氨酰（mCys）残基。这两种蛋白质都含有5个保守的Cys残基，每个残基都是周转所必需的。三个半胱氨酸位于一个典型的RS CxxxCxxC基序和协调的[4Fe-4S]-簇辅因子。其余两个半胱氨酸位于多肽的相对末端。在这里，我们表明，每个蛋白质只包含一个“自由基SAM”[4Fe-4S]集群，和两个剩余的保守的半胱氨酸不协调额外的含铁物种。此外，我们表明，虽然野生型RlmN承担C355 mCys残基在其作为隔离状态，RlmN，无论是工程缺乏[4Fe-4S]簇的协调半胱氨酸的取代，或从大肠杆菌分离培养铁限制条件下，不承担C355 mCys残基。在野生型载脂蛋白RlmN上重建[4Fe-4S]簇，然后加入SAM，导致S-腺苷高半胱氨酸（SAH）和mCys残基的快速产生，而用SAM处理载脂蛋白RlmN没有提供可观察到的反应。这些结果表明，在Cfr和RlmN中，SAM与[4Fe-4S]簇合物的独特铁结合显示两种反应性。它用于甲基化RlmN的C355（Cfr的C338），或用于产生底物依赖性甲基合酶活性所需的5 '-脱氧腺苷5'-自由基。
The radical SAM (RS) proteins RlmN and Cfr catalyze methylation of carbons 2 and 8, respectively, of adenosine 2503 in 23S rRNA. Both reactions are similar in scope, entailing the synthesis of a methyl group partially derived from S-adenosylmethionine (SAM) onto electrophilic sp2-hybridized carbon atoms via the intermediacy of a protein S-methylcysteinyl (mCys) residue. Both proteins contain five conserved Cys residues, each of which is required for turnover. Three cysteines lie in a canonical RS CxxxCxxC motif and coordinate a [4Fe–4S]-cluster cofactor. The remaining two cysteines are at opposite ends of the polypeptide. Herein we show that each protein contains only the one “radical SAM” [4Fe–4S] cluster, and that the two remaining conserved cysteines do not coordinate additional iron-containing species. In addition, we show that while wild-type RlmN bears the C355 mCys residue in its as-isolated state, RlmN that is either engineered to lack the [4Fe–4S] cluster by substitution of the coordinating cysteines, or isolated from Escherichia coli cultured under iron-limiting conditions, does not bear a C355 mCys residue. Reconstitution of the [4Fe–4S] cluster on wild-type apo RlmN followed by addition of SAM results in rapid production of S-adenosylhomocysteine (SAH) and the mCys residue, while treatment of apo RlmN with SAM affords no observable reaction. These results indicate that in Cfr and RlmN, SAM bound to the unique iron of the [4Fe–4S] cluster displays two reactivities. It serves to methylate C355 of RlmN (C338 of Cfr), or it serves to generate the 5’-deoxyadenosyl 5’-radical, required for substrate-dependent methyl synthase activity.
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影响因子：
2.9
作者：
Martinez-Gomez, N. Cecilia;Downs, Diana M.
通讯作者：
Downs, Diana M.
影响因子：
4.9
作者：
Smith, Lisa K.;Mankin, Alexander S.
通讯作者：
Mankin, Alexander S.
自由基 SAM 酶进行甲基转移的结构基础。
DOI：
10.1126/science.1205358
发表时间：
2011-05-27
期刊：
Science (New York, N.Y.)
影响因子：
--
作者：
Boal AK;Grove TL;McLaughlin MI;Yennawar NH;Booker SJ;Rosenzweig AC
通讯作者：
Rosenzweig AC
影响因子：
15
作者：
Yan F;LaMarre JM;Röhrich R;Wiesner J;Jomaa H;Mankin AS;Fujimori DG
通讯作者：
Fujimori DG
影响因子：
7.8
作者：
Booker, Squire J.
通讯作者：
Booker, Squire J.