Liver betaine-hornocysteine S-methyltransferase activity undergoes a redox switch at the active site zinc
Liver betaine-hornocysteine S-methyltransferase activity undergoes a redox switch at the active site zinc
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DOI：
10.1016/j.abb.2008.01.017
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发表时间：
2008-04-01
影响因子：
3.9
通讯作者：
Garrow, Timothy A.
中科院分区：
文献类型：
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作者：
Castro, Carmen;Millian, Norman S.;Garrow, Timothy A.
使用氧化还原惰性的甲基受体，我们表明，甜菜碱-同型半胱氨酸S-甲基转移酶（BHMT）需要一个硫醇还原剂的活动。BHMT短期暴露于无还原剂的缓冲液中可使酶失活，而不会导致其催化锌的任何损失。通过再加入硫醇还原剂可以完全恢复活性。BHMT的催化锌由三个硫醇盐和一个羟基结合。巯基修饰实验表明，当BHMT在无还原剂的缓冲液中无活性时，在三种锌结合配体中的两种之间形成二硫键，并且这种二硫键可以通过重新建立还原条件而容易地还原，同时恢复活性。BHMT长期暴露于无还原剂缓冲液导致其催化Zn的缓慢、不可逆损失和相应的活性损失。使用谷胱甘肽合成严重受损的谷氨酸-半胱氨酸连接酶修饰物亚基敲除小鼠Gclm（-/-）的实验表明，与Gclm（+/+）小鼠相比，Gclm（-/-）小鼠的BHMT活性降低约75%。(c)2008年爱思唯尔公司All rights reserved.
Using a redox-inert methyl acceptor, we show that betaine-homocysteine S-methyltransferase (BHMT) requires a thiol reducing agent for activity. Short-term exposure of BHMT to reducing agent-free buffer inactivates the enzyme without causing any loss of its catalytic zinc. Activity can be completely restored by the re-addition of a thiol reducing agent. The catalytic zinc of BHMT is bound by three thiolates and one hydroxyl group. Thiol modification experiments indicate that a disulfide bond is formed between two of the three zinc-binding ligands when BHMT is inactive in a reducing agent-free buffer, and that this disulfide can be readily reduced with the concomitant restoration of activity by re-establishing reducing conditions. Long-term exposure of BHMT to reducing agent-free buffer results in the slow, irreversible loss of its catalytic Zn and a corresponding loss of activity. Experiments using the glutamate-cysteine ligase modifier subunit knockout mice Gclm(-/-), which are severely impaired in glutathione synthesis, show that BHMT activity is reduced about 75% in Gclm(-/-) compared to Gclm(+/+) mice. (c) 2008 Elsevier Inc. All rights reserved.