Adaptation of an HPLC system for transient-state enzyme kinetic experiments: pulse-flow method.
The components of an HPLC system can be easily reassembled into an instrument for transient-state enzyme kinetic experiments based on the continuous-flow technique. The scheme of reassembly, operation and data evaluation is described in detail for the Pharmacia FPLC system. The working quality of the suggested scheme was tested using a well-studied process of bacterial beta-galactosidase activation by Mg2+ ions. The found parameters were in a good agreement with the literature data. The pulse-flow mode was developed for the delivery of working solutions to reduce the required quantities of enzyme and/or other reaction components. The work presents new approach to HPLC modules as building blocks for sophisticated liquid flow reactors, such as automatic solid phase synthesizers, sequencers and so on.
Tsygankov AYu
A. N. Bakh Institute of Biochemistry, Academy of Sciences of the U.S.S.R., Moscow.
Motorin YuA
Wolfson
A D
AD
Kirpotin
D B
DB
D002851:Chromatography, High Pressure Liquid / Q000295:instrumentation*; D004789:Enzyme Activation; D004798:Enzymes / Q000378:metabolism*; D004867:Equipment Design; D007700:Kinetics; D008274:Magnesium / Q000502:physiology; D011673:Pulsatile Flow; D001616:beta-Galactosidase / Q000378:metabolism
D004798:Enzymes; D001616:beta-Galactosidase; D008274:Magnesium