Characterization of glutamate decarboxylase (GAD) from Lactiplantibacillus plantarum FNCC260 isolated from Indonesian fermented foods
Ida Bagus Agung Yogeswara1*, Dietmar Haltrich2, Thu-Ha Nguyen2

1Department of Nutrition, Universitas Dhyana Pura, Jalan raya Padang Luwih, Kuta utara, Dalung, Bali, Indonesia
2Food Biotechnology Laboratory, Department of Food Science and Technology, University of Natural Resources and Life Science BOKU, Muthgasse, Vienna, Austria

*correspondence email: agungyogeswara[at]undhirabali.ac.id

Abstract

Glutamate decarboxylase (GAD, EC. 4.1.1.15) is a pyridoxal 5^-phosphate (PLP)-dependent enzyme and a key enzyme in decarboxylation of L-glutamate to &#947--aminobutyric acid (GABA) and CO2. 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 it 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 suggesting that GAD from L. plantarum FNCC260 was homodimeric enzyme. The purified GAD has pH and temperature optimum at 4.5 and 60oC respectively. The enzyme showed stability at 4oC and at pH 4.0 with residual activity 94% and 70% respectively. Gradual increase of cofactor PLP did not enhance enzyme activity. GAD activity was enhanced by CaCl2 and MgCl2. Whereas, the activity was inhibited by FeCl3, AgNO3 and CuSO4.

Keywords: glutamate decarboxylase, GABA, L. plantarum, fermented foods

Topic: Food Microbiology and Biotechnology