Production of enzyme reactors after separation by non-denaturing two-dimensional electrophoresis and immobilization on membrane.
Activities of carboxylesterase and malate dehydrogenase on membranes were retained after enzymes of mouse liver cytosol were separated by non-denaturing, two-dimensional electrophoresis (2-DE), stained using imidazole and zinc salts and electroblotted on to membranes. Furthermore, hydrolytic changes of phosphatidylcholine by the esterase were examined using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) after separation, and reversible staining and immobilization to membranes. Hydrolytic activity of the esterase on the membranes was 20% of the original activity of the tissue homogenate. The present method can be applied to the production of several types of enzyme reactors on membranes.
Shimazaki
Youji
Y
Graduate School of Science and Engineering (Science Section) and Venture Business Laboratory, Ehime University, Matsuyama City, 790-8577, Japan. yoji@dpc.ehime-u.ac.jp
Kuroda
Takahiro
T
D000818:Animals; D019149:Bioreactors; D043182:Carboxylesterase / Q000302:isolation & purification*; D004800:Enzymes, Immobilized / Q000378:metabolism*; D008099:Liver / Q000201:enzymology*; D008291:Malate Dehydrogenase / Q000302:isolation & purification*; D008566:Membranes; D051379:Mice
D004800:Enzymes, Immobilized; D008291:Malate Dehydrogenase; D043182:Carboxylesterase