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Determination of lysine decarboxylase activity

KK Kira Küsters
MP Martina Pohl
UK Ulrich Krauss
GÖ Gizem Ölçücü
SA Sandor Albert
KJ Karl-Erich Jaeger
WW Wolfgang Wiechert
MO Marco Oldiges
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Enzyme activity was determined by adding 1 mL of 50 mM Kpi buffer (pH 7.2), 0.1 mM pyridoxalphosphat (PLP) and 10 mM l-lysine to the CatIB pellet, originated from a cell culture with a normalized OD600 nm of 12.5 (See Protein production, cell disruption and protein purification and incubation). The soluble fraction, after cell lysis, was refilled to the normalized volume of 12 mL with Kpi-PLP-l-lysine ratio compared to the CatIB pellet fraction. 1 mL of the solution was used for the enzyme assay. The samples were incubated at 1000 rpm and 30 °C. Samples were taken after 0, 3, 6, 12 and 20 min and the enzyme was inactivated by adding 80 % (v/v) methanol and subsequently l-lysine and DAP concentrations were determined by HPLC to calculate conversion rate.

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