Cloning and expression of rice glutamate decarboxylase (GAD) in Escherichia coli

A. Kariminik; M. Saeidi; P. Parsia

doi:10.14715/cmb/2017.63.9.18

Issue

Vol. 63 No. 9: Issue 9

Issue Published : September 30, 2017

The undersigned hereby assign all rights, included but not limited to copyright, for this manuscript to CMB Association upon its submission for consideration to publication on Cellular and Molecular Biology. The rights assigned include, but are not limited to, the sole and exclusive rights to license, sell, subsequently assign, derive, distribute, display and reproduce this manuscript, in whole or in part, in any format, electronic or otherwise, including those in existence at the time this agreement was signed. The authors hereby warrant that they have not granted or assigned, and shall not grant or assign, the aforementioned rights to any other person, firm, organization, or other entity. All rights are automatically restored to authors if this manuscript is not accepted for publication.

Cloning and expression of rice glutamate decarboxylase (GAD) in Escherichia coli

https://doi.org/10.14715/cmb/2017.63.9.18

A. Kariminik

Department of Microbiology, Kerman Branch, Islamic Azad University, Kerman, Iran

M. Saeidi

Department of Microbiology, Kerman Branch, Islamic Azad University, Kerman, Iran

P. Parsia

Young Researchers and Elite Club, Kerman Branch, Islamic Azad University, Kerman, Iran

Corresponding Author(s) : A. Kariminik

a.kariminik@iauk.ac.ir

Cellular and Molecular Biology, Vol. 63 No. 9: Issue 9
Article Published : September 30, 2017

Abstract

Glutamate decarboxylase (GAD) converts L-glutamate to g-aminobutyric acid (GABA), which is a non-protein amino acid present in all organisms with some activities including improvement of neurve and cytoskeltal functions. Therefore, GAD is considered as a key molecule to use in molecular therapy of genetical human diseases. Accordingly, cloning of GADs from various plants is an important aim of researchers. The aim of this study was to clone rice (Oryza sativa L.) GADs in Escherichia coli (E.coli) MC 1061 bacterium.In this study, rice GADs was cloned in E.coli in both 37°C and 28°C. Two concentrations of Isopropyl-Î²-D-thiogalactoside (IPTG) (0.5mM and 1mM) were investigated in TB medium. Purification was done with Ni2+-nitrilotriacetic acid (NTA) resin in various concentration of imidasol. According to SDS-PAGE analysis, rice GADs was cloned and expressed successfully in E.coli MC 1061 bacterium and the most expression was done in 37°C and 0.5mM IPTG and the best concentration of imidasol was 100mM for purification step. Based on the results, rice GADs can be expressed in E.coli MC 1061 bacterium and, hence, it is a suitable way to produce the plant enzyme in industrial scales.

Keywords

Escherichia coli Glutamate decarboxylase Overexpression.

Kariminik, A., Saeidi, M., & Parsia, P. (2017). Cloning and expression of rice glutamate decarboxylase (GAD) in Escherichia coli. Cellular and Molecular Biology, 63(9), 106–108. https://doi.org/10.14715/cmb/2017.63.9.18

Download Citation

References

O'Byrne CP, Karatzas KA. The role of sigma B (sB) in the stress adaptations of Listeria monocytogenes: overlaps between stress adaptation and virulence. Adv Appl Microbiol. 2008,65(11).
Fait A, Fromm H, Walter D, Galili G, Fernie AR. Highway or byway: the metabolic role of the GABA shunt in plants. Trends Plant Sci. 2008,13(1):14-9. 10.1016/j.tplants.2007.10.005
Shelp BJ, Bown AW, McLean MD. Metabolism and functions of gamma-aminobutyric acid. Trends Plant Sci. 1999,4(11):446-52.
Zhu L, Peng Q, Song F, Jiang Y, Sun C, Zhang J, Huang D. Structure and regulation of the gab gene cluster, involved in the Î³-aminobutyric acid shunt, are controlled by a Ïƒ54 factor in Bacillus thuringiensis. J Bacteriol. 2010,192(1):346-55.
Bouche N, Fromm H. GABA in plants: just a metabolite? Trends Plant Sci. 2004,9(3):110-5.
Bown AW, Macgregor KB, Shelp BJ. Gamma-aminobutyrate: defense against invertebrate pests? Trends Plant Sci. 2006,11(9):424-7. 10.1016/j.tplants.2006.07.002
Akihiro T, Koike S, Tani R, Tominaga T, Watanabe S, Iijima Y, Aoki K, Shibata D, Ashihara H, Matsukura C, Akama K, Fujimura T, Ezura H. Biochemical mechanism on GABA accumulation during fruit development in tomato. Plant Cell Physiol. 2008,49(9):1378-89. 10.1093/pcp/pcn113
Ling V, Snedden WA, Shelp BJ, Assmann SM. Analysis of a soluble calmodulin binding protein from fava bean roots: identification of glutamate decarboxylase as a calmodulin-activated enzyme. Plant Cell. 1994,6(8):1135-43.
Baums G, ChenS Y, Arazis T, Takatsujig H, FrommST H. A Plant Glutamate Decarboxylase Containing a Calmodulin Binding Domain. J Biol Chem. 1993,268:19610-7.
Snedden WA, Arazi T, Fromm H, Shelp BJ. Calcium/Calmodulin Activation of Soybean Glutamate Decarboxylase. Plant Physiol. 1995,108(2):543-9.
Ma S, Huang Y, Yin Z, Menassa R, Brandle JE, Jevnikar AM. Induction of oral tolerance to prevent diabetes with transgenic plants requires glutamic acid decarboxylase (GAD) and IL-4. Proc Natl Acad Sci USA. 2004,101(15):5680-5. 10.1073/pnas.0307420101
Gallego PP, Whotton L, Picton S, Grierson D, Gray JE. A role for glutamate decarboxylase during tomato ripening: the characterisation of a cDNA encoding a putative glutamate decarboxylase with a calmodulin-binding site. Plant Mol Biol. 1995,27(6):1143-51.
Oh SH, Choi WG, Lee IT, Yun SJ. Cloning and characterization of a rice cDNA encoding glutamate decarboxylase. Journal of biochemistry and molecular biology. 2005,38(5):595-601.
Akama K, Akihiro T, Kitagawa M, Takaiwa F. Rice (Oryza sativa) contains a novel isoform of glutamate decarboxylase that lacks an authentic calmodulin-binding domain at the C-terminus. Bba-Gene Struct Expr. 2001,1522(3):143-50. Doi 10.1016/S0167-4781(01)00324-4
Li Z, Wu J, Zhang B, Wang F, Ye X, Huang Y, Huang Q, Cui Z. AmyM, a Novel Maltohexaose-Forming alpha-Amylase from Corallococcus sp. strain EGB. Appl Environ Microbiol. 2015,81(6):1977-87. 10.1128/AEM.03934-14
Calver A, Medhurst A, Robbins M, Charles K, Evans M, Harrison D, Stammers M, Hughes S, Hervieu G, Couve A. The expression of GABA B1 and GABA B2 receptor subunits in the CNS differs from that in peripheral tissues. Neurosci. 2000,100(1):155-70.
Liang C, Li Y, Liu Z, Wu W, Hu B. Protein aggregation formed by recombinant cp19k homologue of Balanus albicostatus combined with an 18 kDa N-terminus encoded by pET-32a (+) plasmid having adhesion strength comparable to several commercial glues. PloS one. 2015,10(8):e0136493.

References

O'Byrne CP, Karatzas KA. The role of sigma B (sB) in the stress adaptations of Listeria monocytogenes: overlaps between stress adaptation and virulence. Adv Appl Microbiol. 2008,65(11).

Fait A, Fromm H, Walter D, Galili G, Fernie AR. Highway or byway: the metabolic role of the GABA shunt in plants. Trends Plant Sci. 2008,13(1):14-9. 10.1016/j.tplants.2007.10.005

Shelp BJ, Bown AW, McLean MD. Metabolism and functions of gamma-aminobutyric acid. Trends Plant Sci. 1999,4(11):446-52.

Zhu L, Peng Q, Song F, Jiang Y, Sun C, Zhang J, Huang D. Structure and regulation of the gab gene cluster, involved in the Î³-aminobutyric acid shunt, are controlled by a Ïƒ54 factor in Bacillus thuringiensis. J Bacteriol. 2010,192(1):346-55.

Bouche N, Fromm H. GABA in plants: just a metabolite? Trends Plant Sci. 2004,9(3):110-5.

Bown AW, Macgregor KB, Shelp BJ. Gamma-aminobutyrate: defense against invertebrate pests? Trends Plant Sci. 2006,11(9):424-7. 10.1016/j.tplants.2006.07.002

Akihiro T, Koike S, Tani R, Tominaga T, Watanabe S, Iijima Y, Aoki K, Shibata D, Ashihara H, Matsukura C, Akama K, Fujimura T, Ezura H. Biochemical mechanism on GABA accumulation during fruit development in tomato. Plant Cell Physiol. 2008,49(9):1378-89. 10.1093/pcp/pcn113

Ling V, Snedden WA, Shelp BJ, Assmann SM. Analysis of a soluble calmodulin binding protein from fava bean roots: identification of glutamate decarboxylase as a calmodulin-activated enzyme. Plant Cell. 1994,6(8):1135-43.

Baums G, ChenS Y, Arazis T, Takatsujig H, FrommST H. A Plant Glutamate Decarboxylase Containing a Calmodulin Binding Domain. J Biol Chem. 1993,268:19610-7.

Snedden WA, Arazi T, Fromm H, Shelp BJ. Calcium/Calmodulin Activation of Soybean Glutamate Decarboxylase. Plant Physiol. 1995,108(2):543-9.

Ma S, Huang Y, Yin Z, Menassa R, Brandle JE, Jevnikar AM. Induction of oral tolerance to prevent diabetes with transgenic plants requires glutamic acid decarboxylase (GAD) and IL-4. Proc Natl Acad Sci USA. 2004,101(15):5680-5. 10.1073/pnas.0307420101

Gallego PP, Whotton L, Picton S, Grierson D, Gray JE. A role for glutamate decarboxylase during tomato ripening: the characterisation of a cDNA encoding a putative glutamate decarboxylase with a calmodulin-binding site. Plant Mol Biol. 1995,27(6):1143-51.

Oh SH, Choi WG, Lee IT, Yun SJ. Cloning and characterization of a rice cDNA encoding glutamate decarboxylase. Journal of biochemistry and molecular biology. 2005,38(5):595-601.

Akama K, Akihiro T, Kitagawa M, Takaiwa F. Rice (Oryza sativa) contains a novel isoform of glutamate decarboxylase that lacks an authentic calmodulin-binding domain at the C-terminus. Bba-Gene Struct Expr. 2001,1522(3):143-50. Doi 10.1016/S0167-4781(01)00324-4

Li Z, Wu J, Zhang B, Wang F, Ye X, Huang Y, Huang Q, Cui Z. AmyM, a Novel Maltohexaose-Forming alpha-Amylase from Corallococcus sp. strain EGB. Appl Environ Microbiol. 2015,81(6):1977-87. 10.1128/AEM.03934-14

Calver A, Medhurst A, Robbins M, Charles K, Evans M, Harrison D, Stammers M, Hughes S, Hervieu G, Couve A. The expression of GABA B1 and GABA B2 receptor subunits in the CNS differs from that in peripheral tissues. Neurosci. 2000,100(1):155-70.

Liang C, Li Y, Liu Z, Wu W, Hu B. Protein aggregation formed by recombinant cp19k homologue of Balanus albicostatus combined with an 18 kDa N-terminus encoded by pET-32a (+) plasmid having adhesion strength comparable to several commercial glues. PloS one. 2015,10(8):e0136493.

Author biographies is not available.