A continuous kinetic assay for adenylation enzyme activity and inhibition.
A continuous kinetic assay for adenylation enzyme activity and inhibition.
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DOI：
10.1016/j.ab.2010.04.033
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发表时间：
2010-09-01
影响因子：
2.9
通讯作者：
Aldrich CC
中科院分区：
文献类型：
--
作者：
Wilson DJ;Aldrich CC
Adenylation/adenylate-forming enzymes catalyze the activation of a carboxylic acid at the expense of ATP to form an acyl-adenylate intermediate and pyrophosphate (PPi). In a second half-reaction, adenylation enzymes catalyze the transfer of the acyl moiety of the acyl-adenylate onto an acceptor molecule, which can be either a protein or a small molecule. We describe the design, development, and validation of a coupled continuous spectrophotometric assay for adenylation enzymes that employs hydroxylamine as a surrogate acceptor molecule leading to the formation of a hydroxamate. The released pyrophosphate from the first half-reaction is measured using the pyrophosphatase–purine nucleoside phosphorylase coupling system with the chromogenic substrate 7-methylthioguanosine (MesG). The coupled hydroxamate–MesG assay is especially useful for characterizing the activity and inhibition of adenylation enzymes that acylate a protein substrate and/or fail to undergo rapid ATP-PPi exchange.
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影响因子：
14.8
作者：
Arora, Pooja;Goyal, Aneesh;Natarajan, Vivek T.;Rajakumara, Eerappa;Verma, Priyanka;Gupta, Radhika;Yousuf, Malikmohamed;Trivedi, Omita A.;Mohanty, Debasisa;Tyagi, Anil;Sankaranarayanan, Rajan;Gokhale, Rajesh S.
通讯作者：
Gokhale, Rajesh S.
影响因子：
4.8
作者：
STACHELHAUS, T;MARAHIEL, MA
通讯作者：
MARAHIEL, MA
影响因子：
2.9
作者：
Quadri, LEN;Weinreb, PH;Walsh, CT
通讯作者：
Walsh, CT
影响因子：
2.9
作者：
Luo, LS;Walsh, CT
通讯作者：
Walsh, CT
影响因子：
7.3
作者：
Somu, Ravindranadh V.;Wilson, Daniel J.;Aldrich, Courtney C.
通讯作者：
Aldrich, Courtney C.