Calcium/calmodulin activation of two divergent glutamate decarboxylases from tobacco.
Glutamate decarboxylase (GAD, EC 4.1.1.15) catalyses the alpha-decarboxylation of glutamate to produce gamma-aminobutyrate (GABA). The nucleotide sequences of two divergent GADs (designated GAD1 and GAD3) were isolated from a Nicotiana tabacum L. cv. Samsun NN leaf cDNA library. Open reading frames indicated that GAD1 encodes a polypeptide of 496 amino acids and has greater than 99% identity with known tobacco GADs, whereas GAD3 encodes a polypeptide of 491 amino acids and has about 14% divergence from known tobacco GADs. Genomic DNA analysis suggested that there are at least four tobacco GAD genes, existing in pairs of highly identical genes. An in vitro assay at pH 7.3 revealed that activities of the recombinant proteins, after isolation from Escherichia coli and partial purification by nickel-affinity chromatography, are 57-133 times the control levels in the presence of 0.5 mM calcium and 0.2 micro M bovine calmodulin.
Yevtushenko
Dmytro P
DP
Department of Plant Agriculture, Biotechnology Division, Bovey Building, University of Guelph, Guelph, Ontario, Canada N1G 2W1.
McLean
Michael D
MD
Peiris
Sriyani
S
Van Cauwenberghe
Owen R
OR
Shelp
Barry J
BJ
D002118:Calcium / Q000494:pharmacology*; D002147:Calmodulin / Q000494:pharmacology*; D003001:Cloning, Molecular; D004789:Enzyme Activation / Q000187:drug effects; D017343:Genes, Plant / Q000235:genetics; D005968:Glutamate Decarboxylase / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D014026:Nicotiana / Q000187:drug effects* / Q000201:enzymology* / Q000235:genetics
D002147:Calmodulin; D020033:Protein Isoforms; D005968:Glutamate Decarboxylase; D002118:Calcium