Liver betaine-homocysteine S-methyltransferase activity undergoes a redox switch at the active site zinc

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...

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Bibliographic Details
Published in:Archives of biochemistry and biophysics 2008-04, Vol.472 (1), p.26-33
Main Authors: Castro, Carmen, Millian, Norman S., Garrow, Timothy A.
Format: Article
Language:English
Subjects:
Betaine-Homocysteine S-Methyltransferase - chemistry
BHMT
Binding Sites
Catalysis
Cysteine
Enzyme Activation
Glutamate-cysteine ligase modifier subunit Gclm(−/−) knockout mouse
Glutathione
Humans
Liver - enzymology
Methionine
Oxidation-Reduction
Oxidative stress
Protein Binding
Sulfhydryl Compounds - chemistry
Zinc - chemistry
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Summary: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.
ISSN:0003-9861
1096-0384
DOI:10.1016/j.abb.2008.01.017