Journal
ARCHIVES OF BIOCHEMISTRY AND BIOPHYSICS
Volume 472, Issue 1, Pages 26-33Publisher
ELSEVIER SCIENCE INC
DOI: 10.1016/j.abb.2008.01.017
Keywords
BHMT; cysteine; glutathione; methionine; glutamate-cysteine ligase modifier subunit Gclm(-/-) knockout mouse; oxidative stress
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Funding
- NIDDK NIH HHS [DK52501, R29 DK052501, R01 DK052501-09, R01 DK052501] Funding Source: Medline
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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.
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