A residue outside the active site CXXC motif regulates the catalytic efficiency of Glutaredoxin 3.
A residue outside the active site CXXC motif regulates the catalytic efficiency of Glutaredoxin 3.
复制标题
DOI：
10.1039/b912753d
复制
发表时间：
2010-01
影响因子：
--
通讯作者：
Keinan E
中科院分区：
文献类型：
--
作者：
Shekhter T;Metanis N;Dawson PE;Keinan E
The glutaredoxin (Grx) family of oxidoreductases has a conserved residue at position 8 that varies between Arginine in Grx1 and Lysine in Grx3. It has been proposed that this Arg/Lys change is the main cause for the 35 mV difference in redox potential between the two enzymes. To gain insights into the catalytic machinery of Grx3 and directly evaluate the role of residue 8 in the catalysis of thiol-disulfide exchange by this enzyme, we synthesized the “wild type” enzyme (sGrx3), and four analogues substituting the lysine at position 8 with arginine, ornithine (Orn), citrulline (Cit) and norvaline (Nva). The redox potential and equilibration kinetics with thioredoxin (Trx1) were determined for each enzyme by fluorescence intensity. While minor effects on redox potential were observed, we found that residue 8 had a more marked effect on the catalytic efficiency of this enzyme. Surprisingly, truncation of the functional group resulted in a more efficient enzyme, Lys8Nva, exhibiting rate constants that are an order of magnitude higher than sGrx3 for both forward and reverse reactions. These observations pose the question why would a residue that reduces the rate of enzyme turnover be evolutionarily conserved? The significant changes in the kinetic parameters suggest that this position plays an important role in the thiol-disulfide exchange reaction by affecting the nucleophilic thiolate through electrostatic or hydrogen bonding interactions. Since the reduced Grx has an exposed thiol that could easily be alkylated, either Arg or Lys could act as a gatekeeper that deters unwanted electrophiles from attacking the active site thiolate.
登录
查看更多内容
影响因子：
56.9
作者：
DAWSON, PE;MUIR, TW;KENT, SBH
通讯作者：
KENT, SBH
影响因子：
5.6
作者：
Foloppe, N;Sagemark, J;Nilsson, L
通讯作者：
Nilsson, L
DOI：
10.1073/pnas.93.14.6851
发表时间：
1996-07-09
影响因子：
11.1
作者：
Fitzgerald, MC;Chernushevich, I;Kent, SBH
通讯作者：
Kent, SBH
影响因子：
2.9
作者：
Jao, SC;Ospina, SME;Mieyal, JJ
通讯作者：
Mieyal, JJ
影响因子：
4.6
作者：
ARNOLD, AP;TAN, KS;RABENSTEIN, DL
通讯作者：
RABENSTEIN, DL