Anaerobic sulfatase-maturating enzyme--a mechanistic link with glycyl radical-activating enzymes?
Anaerobic sulfatase-maturating enzyme--a mechanistic link with glycyl radical-activating enzymes?
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DOI：
10.1111/j.1742-4658.2010.07613.x
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发表时间：
2010-04
期刊：
影响因子：
--
通讯作者：
Berteau O
中科院分区：
文献类型：
--
作者：
Benjdia A;Subramanian S;Leprince J;Vaudry H;Johnson MK;Berteau O
硫酸盐酶是存在于原核生物和真核生物中的一大类酶。这类水解酶的独特之处在于需要对关键活性位点的半胱氨酸或丝氨酸残基进行必要的翻译后修饰，使其成为c - α-甲酰基甘氨酸（FGly）。在此，我们报道了一种独特的自由基adomet酶的机制研究，anSMEs（厌氧硫酸酯酶成熟酶）催化cys型和ser型硫酸酯酶的氧化，并具有三个[4Fe-4S]2+，+簇。我们能够开发出一种可靠的定量酶分析方法，可以直接测量FGly的产生和AdoMet的裂解。结果表明AdoMet切割和FGly形成的化学计量偶联使用肽底物与半胱氨酸或丝氨酸活性位点残基。重组的野生型酶和半胱氨酸簇突变体的分析和EPR研究表明，存在三个几乎等电位的[4Fe-4S]2+，+簇，每一个都是体外FGly生成所必需的。更令人惊讶的是，我们的数据表明，两个额外的[4Fe-4S]2+，+簇是AdoMet获得有效还原切割所必需的，这表明它们参与了AdoMet [4Fe-4S]2+，+中心的还原。这些结果，再加上最近anSMEs使用5 ' -脱氧腺苷基自由基直接从活性位点半胱氨酸或丝氨酸残基中提取Cβ h原子的证明，为这类新的自由基- adomet酶的机制提供了新的见解。
Sulfatases form a major group of enzymes present in prokaryotes and eukaryotes. This class of hydrolases is unique in requiring an essential post-translational modification of a critical active-site cysteinyl or seryl residue to Cα-formylglycine (FGly). Herein, we report mechanistic investigations of a unique class of radical-AdoMet enzymes, anSMEs (anaerobic sulfatase-maturating enzymes) which catalyze the oxidation of Cys-type and Ser-type sulfatases and possess three [4Fe-4S]2+,+ clusters. We were able to develop a reliable quantitative enzymatic assay which allowed the direct measurement of FGly production and AdoMet cleavage. The results demonstrate stoichiometric coupling of AdoMet cleavage and FGly formation using peptide substrates with cysteinyl or seryl active-site residues. Analytical and EPR studies of the reconstituted wild-type enzyme and cysteinyl cluster mutants indicate the presence of three almost isopotential [4Fe-4S]2+,+ clusters, each of which is required for in vitro FGly generation. More surprisingly, our data indicate that the two additional [4Fe-4S]2+,+ clusters are required to obtain efficient reductive cleavage of AdoMet suggesting their involvement in the reduction of the radical AdoMet [4Fe-4S]2+,+ center. These results, in addition to the recent demonstration of direct abstraction by anSMEs of the Cβ H-atom from the active site cysteinyl or seryl residue using a 5′-deoxyadenosyl radical, provide new insights into the mechanism of this new class of radical-AdoMet enzymes.
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影响因子：
15
作者：
Chen, DW;Walsby, C;Frey, PA
通讯作者：
Frey, PA
影响因子：
4.9
作者：
Friedel, MG;Berteau, O;Carell, T
通讯作者：
Carell, T
影响因子：
3.1
作者：
Hoffman, JA;Badger, JL;Kim, KS
通讯作者：
Kim, KS
DOI：
10.1073/pnas.0510501103
发表时间：
2006-05-16
影响因子：
11.1
作者：
Hagelueken, Gregor;Adams, Thorsten M.;Schubert, Wolf-Dieter
通讯作者：
Schubert, Wolf-Dieter
影响因子：
15
作者：
Benjdia, Alhosna;Leprince, Jerome;Berteau, Olivier
通讯作者：
Berteau, Olivier