• Home
  • Search Results
  • Solubilization of a membrane-bound diol dehydratase with retention of EPR g = 2.02 signal by using 2-(N-cyclohexylamino)ethanesulfonic acid buffer.

Solubilization of a membrane-bound diol dehydratase with retention of EPR g = 2.02 signal by using 2-(N-cyclohexylamino)ethanesulfonic acid buffer.

Proceedings of the National Academy of Sciences of the United States of America (1987-01-01)
M G Hartmanis, T C Stadtman
ABSTRACT

A procedure for solubilization of the oxygen-labile, membrane-bound diol dehydratase from Clostridium glycolicum with retention of enzymatic activity is described. The procedure involves sonication of crude membrane preparations anaerobically in 0.1 M 2-(N-cyclohexylamino)ethane-sulfonic acid (CHES) buffer (pH 8.6-9.0) containing 2 mM dithiothreitol. The addition of dimethylsulfoxide (30%) and lysophosphatidylcholine (0.15 mg/ml) to the solubilization buffer resulted in a 10-fold increase in recovery of solubilized diol dehydratase activity. After ultracentrifugation, an overall recovery of 50% of the activity initially present in the crude membrane preparations was achieved. Active membrane preparations and the solubilized enzyme exhibited an EPR signal at g = 2.02. Both enzyme activity and EPR signal were sensitive to oxygen and the radical scavengers, NH2OH and hydroxy-urea.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
CHES, ≥99.0% (titration)
Sigma-Aldrich
CHES, BioXtra, ≥99.0% (titration)
Sigma-Aldrich
CHES, BioUltra, ≥99.5% (T)

Social Media

LinkedIn icon
Twitter icon
Facebook Icon
Instagram Icon

MilliporeSigma

Research. Development. Production.

We are a leading supplier to the global Life Science industry with solutions and services for research, biotechnology development and production, and pharmaceutical drug therapy development and production.

© 2021 Merck KGaA, Darmstadt, Germany and/or its affiliates. All Rights Reserved.

Reproduction of any materials from the site is strictly forbidden without permission.