title: Expression, purification, and characterization of recombinant mangrove glutamine synthetase.
abstract: To expand our knowledge about the relationship of nitrogen use efficiency and glutamine synthetase (GS) activity in the mangrove plant, a cytosolic GS gene from Avicennia marina has been heterologously expressed in and purified from Escherichia coli. Synthesis of the mangrove GS enzyme in E. coli was demonstrated by functional genetic complementation of a GS deficient mutant. The subunit molecular mass of GSI was ~40 kDa. Optimal conditions for biosynthetic activity were found to be 35 °C at pH 7.5. The Mg(2+)-dependent biosynthetic activity was strongly inhibited by Ni(2+), Zn(2+), and Al(3+), whereas was enhanced by Co(2+). The apparent K m values of AmGLN1 for the substrates in the biosynthetic assay were 3.15 mM for glutamate, and 2.54 mM for ATP, 2.80 mM for NH4 (+) respectively. The low affinity kinetics of AmGLN1 apparently participates in glutamine synthesis under the ammonium excess conditions.
authors.affiliation: Shanghai Key Laboratory of Agricultural Genetics and Breeding, Biotechnology Research Institute, Shanghai Academy of Agricultural Sciences, 2901 Beidi Road, Shanghai, People's Republic of China.
authors.forename: Wei
authors.initials: W
authors.lastname: Zhao
authors.forename: Jun
authors.initials: J
authors.lastname: Yang
authors.forename: Yongsheng
authors.initials: Y
authors.lastname: Tian
authors.forename: Xiaoyan
authors.initials: X
authors.lastname: Fu
authors.forename: Bo
authors.initials: B
authors.lastname: Zhu
authors.forename: Yong
authors.initials: Y
authors.lastname: Xue
authors.forename: Jianjie
authors.initials: J
authors.lastname: Gao
authors.forename: Hong-Juan
authors.initials: HJ
authors.lastname: Han
authors.forename: Rihe
authors.initials: R
authors.lastname: Peng
authors.forename: Quan-Hong
authors.initials: QH
authors.lastname: Yao
mesh_terms: D000255:Adenosine Triphosphate / Q000096:biosynthesis; D000641:Ammonia / Q000378:metabolism; D032407:Avicennia / Q000201:enzymology*; D004926:Escherichia coli; D005816:Genetic Complementation Test; D005974:Glutamate-Ammonia Ligase / Q000235:genetics* / Q000378:metabolism*; D018698:Glutamic Acid / Q000096:biosynthesis; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D019216:Metals, Heavy / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics* / Q000378:metabolism*; D013696:Temperature
chemical_list: D019216:Metals, Heavy; D011994:Recombinant Proteins; D018698:Glutamic Acid; D000641:Ammonia; D000255:Adenosine Triphosphate; D005974:Glutamate-Ammonia Ligase
references.citation: Plant Physiol. 1985 Apr;77(4):791-4
references.pmid: 16664139
references.citation: Nucleic Acids Res. 2004 Jul 07;32(12):e98
references.pmid: 15240836
references.citation: Mol Syst Biol. 2006;2:2006.0008
references.pmid: 16738554
references.citation: Phytochemistry. 2008 Jan;69(2):382-9
references.pmid: 17888468
references.citation: Tree Physiol. 2010 Sep;30(9):1148-60
references.pmid: 20566581
references.citation: Mol Gen Genet. 1991 Nov;230(1-2):145-54
references.pmid: 1684022
references.citation: J Biol Chem. 1996 Nov 22;271(47):29561-8
references.pmid: 8939884
references.citation: J Exp Bot. 2011 Feb;62(4):1375-90
references.pmid: 20959627
references.citation: Plant Cell Physiol. 2004 Nov;45(11):1640-7
references.pmid: 15574840
references.citation: Plant Cell Physiol. 2002 Jul;43(7):802-9
references.pmid: 12154143
references.citation: New Phytol. 2005 Jul;167(1):31-9
references.pmid: 15948827
references.citation: J Exp Bot. 2005 Feb;56(412):663-71
references.pmid: 15642719
references.citation: Proc Natl Acad Sci U S A. 1993 Apr 1;90(7):3009-13
references.pmid: 8096645
references.citation: Eur J Biochem. 1990 Oct 24;193(2):319-24
references.pmid: 1977583
references.citation: Science. 1986 Jun 6;232(4755):1242-4
references.pmid: 2871626
references.citation: J Agric Food Chem. 2006 Feb 8;54(3):796-802
references.pmid: 16448185
references.citation: Plant Cell. 2006 Nov;18(11):3252-74
references.pmid: 17138698
references.citation: Mol Syst Biol. 2011 Oct 11;7:539
references.pmid: 21988835
references.citation: Plant Physiol. 1996 Nov;112(3):987-96
references.pmid: 8938407
references.citation: Plant Cell Physiol. 2006 Feb;47(2):299-303
references.pmid: 16338958
references.citation: J Biol Chem. 2004 Apr 16;279(16):16598-605
references.pmid: 14757761
references.citation: J Med Chem. 2005 Oct 6;48(20):6340-9
references.pmid: 16190760
references.citation: Tree Physiol. 2004 Jul;24(7):729-36
references.pmid: 15123444
references.citation: J Exp Bot. 2002 Apr;53(370):979-87
references.pmid: 11912240
references.citation: J Biol Chem. 2006 Sep 29;281(39):29287-96
references.pmid: 16829528
references.citation: Plant Physiol. 1980 Oct;66(4):619-23
references.pmid: 16661490
references.citation: Mol Biotechnol. 2008 May;39(1):49-56
references.pmid: 18074244
references.citation: Protein Sci. 1997 Dec;6(12):2617-23
references.pmid: 9416610
references.citation: Fungal Genet Biol. 2008 Jul;45(7):1103-10
references.pmid: 18479949
references.citation: Biochemistry. 1966 Mar;5(3):872-9
references.pmid: 5911299
references.citation: BMC Plant Biol. 2010 Aug 19;10:183
references.pmid: 20723225
references.citation: J Exp Bot. 2007;58(9):2319-27
references.pmid: 17350935
references.citation: Biometals. 2001 Mar;14(1):13-22
references.pmid: 11368271
references.citation: Sci Rep. 2012;2:419
references.pmid: 22639727