Characterization of glutamate decarboxylase (GAD) from Lactiplantibacillus plantarum FNCC260 isolated from Indonesian fermented foods

• Main Authors: Yogeswara, Ida Bagus Agung, Haltrich, Dietmar, Nguyen, Thu-Ha
• Format: Conference Proceeding
• Language: English
• Subjects: Calcium chloride; E coli; Enzyme activity; Enzymes; Eukaryotes; Ferric chloride; Magnesium chloride; Prokaryotes; Silver nitrate
• ISSN: 0094-243X; 1551-7616
• DOI: 10.1063/5.0119072

Glutamate decarboxylase is a key enzyme in synthesizing the γ-aminobutyric acid (GABA) which occurred by binding with Pyridoxal 5′-Phosphate (PLP). GAD is ubiquitously found in eukaryotes and prokaryotes. The gadB gene encoding glutamate decarboxylase was amplified by PCR, resulting in 1410 bp. The gadB gene was cloned to the expression vector pET 21a (+) and transformed to E. coli T7 harboring pGRO7. The gadB was overexpressed by inducing 0.5 mM IPTG, and recombinant protein was purified using Ion-Metal Affinity Chromatography (IMAC). The apparent molecular masses as judged by SDS-PAGE was 51 kDa. Confirmation of the molecular masses using LC-ESI-MS suggests that GAD from L. plantarum FNCC260 was a homodimeric enzyme. The purified GAD has pH and temperature optimum at 4.5 and 60 °C, respectively. The enzyme showed stability at 4 °C and pH 4.0 with 94% and 70% residual activity. Gradual increase of cofactor PLP did not enhance enzyme activity. CaCl2 and MgCl2 enhanced GAD activity. In contrast, the activity was inhibited by FeCl3, AgNO3, and CuSO4.