Measurement of delta-(L-alpha-aminoadipyl)-L-cysteinyl-D-valine synthetase activity in Streptomyces clavuligerus by high-performance liquid chromatography after precolumn derivatization with o-phthaldialdehyde.
A procedure for rapid, sensitive measurement of delta-(L-alpha-aminoadipyl)-L-cysteinyl-D-valine (ACV) in complex cell extracts has been developed by adapting a widely used method for high-performance liquid chromatography analysis of amino acids. Samples were oxidized with performic acid and derivatized with o-phthaldialdehyde-mercaptoethanol to give the fluorescent isoindole derivative of the peptide sulfonate. The procedure was used to assay Streptomyces clavuligerus cell extracts and partially purified fractions for ACV synthetase activity and to determine some characteristics of the enzyme reaction. The presence of a second enzyme with cysteine as a substrate was also indicated.
White
R L
RL
Chemistry Department, Acadia University, Wolfville, Nova Scotia, Canada.
DeMarco
A C
AC
Shapiro
S
S
Vining
L C
LC
Wolfe
S
S
D002851:Chromatography, High Pressure Liquid; D007202:Indicators and Reagents; D010084:Oxidation-Reduction; D010453:Peptide Synthases / Q000378:metabolism*; D013302:Streptomyces / Q000201:enzymology*; D009764:o-Phthalaldehyde
D007202:Indicators and Reagents; D009764:o-Phthalaldehyde; D010453:Peptide Synthases; C095368:alpha-aminoadipyl-cysteinyl-valine synthetase