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Measurement of glyoxalase activities
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Arai, Makoto, Nihonmatsu‑Kikuchi, Naomi, Itokawa, Masanari, Rabbani, Naila and Thornalley, Paul J. (2014) Measurement of glyoxalase activities. Biochemical Society Transactions, 42 (2). pp. 491-494. 491. doi:10.1042/BST20140010
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Official URL: http://dx.doi.org/10.1042/BST20140010
Abstract
Glyoxalase I catalyses the isomerization of the hemithioacetal formed non-enzymatically from methylglyoxal and glutathione to S-D-lactoylglutathione. The activity of glyoxalase I is conventionally measured spectrophotometrically by following the increase in A240 for which the change in molar absorption coefficient Δε240=2.86 mM⁻¹·cm⁻¹. The hemithioacetal is pre-formed in situ by incubation of methylglyoxal and glutathione in 50 mM sodium phosphate buffer (pH 6.6) at 37°C for 10 min. The cell extract is then added, the A240 is monitored over 5 min, and the initial rate of increase in A240 and hence glyoxalase I activity deduced with correction for blank. Glyoxalase I activity is given in units per mg of protein or cell number where one unit is the amount of enzyme that catalyses the formation of 1 μmol of S-D-lactoylglutathione per min under assay conditions. Glyoxalase II catalyses the hydrolysis of S-D-lactoylglutathione to D-lactate and glutathione. Glyoxalase II activity is also measured spectrophotometrically by following the decrease in A240 for which the change in molar absorption coefficient Δε240=-3.10 mM⁻¹·cm⁻¹. It is given in units per mg of protein or cell number where one unit is the amount of enzyme that catalyses the hydrolysis of 1 μmol of S-D-lactoylglutathione per min under assay conditions. Glyoxalase I and glyoxalase II activity measurements have been modified for use with a UV-transparent microplate for higher sample throughput.
| Item Type: | Journal Article | ||||||
|---|---|---|---|---|---|---|---|
| Subjects: | Q Science > QR Microbiology | ||||||
| Divisions: | Faculty of Medicine > Warwick Medical School > Health Sciences Faculty of Medicine > Warwick Medical School |
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| Library of Congress Subject Headings (LCSH): | Glyoxalase, Gene expression | ||||||
| Journal or Publication Title: | Biochemical Society Transactions | ||||||
| Publisher: | Portland Press Ltd. | ||||||
| ISSN: | 0300-5127 | ||||||
| Official Date: | 1 April 2014 | ||||||
| Dates: |
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| Volume: | 42 | ||||||
| Number: | 2 | ||||||
| Number of Pages: | 4 | ||||||
| Page Range: | pp. 491-494 | ||||||
| Article Number: | 491 | ||||||
| DOI: | 10.1042/BST20140010 | ||||||
| Status: | Peer Reviewed | ||||||
| Publication Status: | Published | ||||||
| Access rights to Published version: | Restricted or Subscription Access | ||||||
| Funder: | Nihon Gakujutsu Shinkōkai [Japan Society for the Promotion of Science] (NGS) |
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