A novel mechanism of V-type zinc inhibition of glutamate dehydrogenase results from disruption of subunit interactions necessary for efficient catalysis.
A novel mechanism of V-type zinc inhibition of glutamate dehydrogenase results from disruption of subunit interactions necessary for efficient catalysis.
复制标题
V 型锌抑制谷氨酸脱氢酶的一种新机制是由于有效催化所需的亚基相互作用被破坏所致。
DOI：
10.1111/j.1742-4658.2011.08240.x
复制
发表时间：
2011-09
期刊：
影响因子：
--
通讯作者：
Bell E
中科院分区：
文献类型：
--
作者：
Bailey J;Powell L;Sinanan L;Neal J;Li M;Smith T;Bell E
锌可有效抑制牛谷氨酸脱氢酶，主要影响Vmax，表明对催化或产物释放具有V型效应。锌抑制降低谷氨酸浓度降低，表明亚基相互作用的作用。与一元羧酸氨基酸，正缬氨酸，这给出了没有证据的亚基相互作用锌不抑制。锌显著降低反应中的预稳态爆发的大小，但不影响酶-NADPH-谷氨酸盐复合物中的NADPH结合，其再次控制稳态转换，表明锌破坏催化能力所需的亚基相互作用。虽然差示扫描量热法表明锌结合并诱导蛋白质的略微构象上更刚性的状态，但有限的蛋白质水解表明天线区域附近的区域和三聚体-三聚体界面变得更加灵活。与锌和铕结合的GDH的结构表明，锌在六聚体中的亚基的三个二聚体之间结合，该区域显示结合阻断催化周转的新型抑制剂，并且与上述发现一致。相反，铕结合到天线区域的基础，似乎废除锌的抑制作用。各种状态的酶的结构已经表明，这两个区域都大量参与与催化营业额相关的构象变化。这些结果表明，V型抑制谷氨酸作为底物产生的结果从破坏亚基相互作用所必需的有效催化，而不是通过对活性位点构象的直接影响。
Bovine Glutamate Dehydrogenase is potently inhibited by zinc and the major impact is on Vmax suggesting a V-type effect on catalysis or product release. Zinc inhibition decreases as glutamate concentrations decrease suggesting a role for subunit interactions. With the monocarboxylic amino acid, norvaline, which gives no evidence of subunit interactions zinc does not inhibit. Zinc significantly decreases the size of the pre-steady state burst in the reaction but does not affect NADPH binding in the Enzyme-NADPH-glutamate complex that governs the steady state turn-over again suggesting that zinc disrupts subunit interactions required for catalytic competence. While differential scanning calorimetry suggests zinc binds and induces a slightly conformationally more rigid state of the protein, limited proteolysis indicate regions in the vicinity of the antennae regions and the trimer-trimer interface become more flexible. The structures of GDH bound with zinc and europium show that zinc binds between the three dimers of subunits in the hexamer, a region shown to bind novel inhibitors that block catalytic turnover and is consistent with the above findings. In contrast, europium binds to the base of the antenna region and appears to abrogate the inhibitory effect of zinc. Structures of various states of the enzyme have shown that both regions are heavily involved in the conformational changes associated with catalytic turnover. These results suggest that the V-type inhibition produced with glutamate as the substrate results from disruption of subunit interactions necessary for efficient catalysis rather than by a direct effect on the active site conformation.
登录
查看更多内容
DOI：
10.1107/s0907444904019158
发表时间：
2004-12-01
影响因子：
2.2
作者：
Emsley, P;Cowtan, K
通讯作者：
Cowtan, K
影响因子：
2.9
作者：
Banerjee, S;Schmidt, T;Smith, TJ
通讯作者：
Smith, TJ
影响因子：
4.1
作者：
ENGEL, PC;DALZIEL, K
通讯作者：
DALZIEL, K
影响因子：
4.1
作者：
ALEX, S;BELL, JE
通讯作者：
BELL, JE
影响因子：
2.9
作者：
BANERJEE, A;LEVY, HR;BELL, JE
通讯作者：
BELL, JE