Anion-Exchange Analysis of Ribulose-Bisphosphate Carboxylase/Oxygenase Reactions: CO2/O2 Specificity Determination and Identification of Side Products
✍ Scribed by M.R. Harpel; E.H. Lee; F.C. Hartman
- Publisher
- Elsevier Science
- Year
- 1993
- Tongue
- English
- Weight
- 659 KB
- Volume
- 209
- Category
- Article
- ISSN
- 0003-2697
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✦ Synopsis
An improved anion-exchange chromatographic method for determining the carboxylation/oxygenation specificity (tau) of ribulose 1,5-bisphosphate carboxylase/oxygenase is presented. This assay, which entails radiometric detection of [1-3H]ribulose-bisphosphate turnover products separated on MonoQ anion-exchange resin, is more convenient, less error-prone, and more generally applicable than previous methods of tau determination. It is suitable for both wild-type and site-directed mutant enzymes of widely varying activity and specificity levels and allows simultaneous visualization of various side products of ribulose-bisphosphate processing. A facile method for scrubbing dissolved O2 from carboxylase reaction solutions, which does not require extensive purging and exchange of dissolved gases, is also described.