Premium
Molecular characterization of the 1‐Deoxy‐D‐xylulose‐5‐ phosphate Synthase (DXS) from marine bacterium, Kocuria gwangalliensis
Author(s) -
kim young tae
Publication year - 2012
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.26.1_supplement.981.2
Subject(s) - recombinant dna , fusion protein , biology , atp synthase , plasmid , microbiology and biotechnology , biochemistry , lac operon , gene , bacteria , expression vector , escherichia coli , genetics
In addition to the ubiquitous mevalonate pathway, Kocuria gwangalliensis utilizes the nonmevalonate pathway for isopentenyl diphosphate biosynthesis. 2‐C‐methyl‐D‐erythritol‐4‐phosphate synthase (MEP synthase) catalyzes the rearrangement of 1‐D‐deoxyxylulose‐ 5‐phosphate (DXP) to methylerythritol‐4‐ phosphate(MEP) as the first pathway‐specific reaction in the MEP biosynthetic pathway. In the present study, the gene coding for 1‐ Deoxy‐D‐xylulose‐5‐phosphate Synthase (DXS) was isolated from the marine bacterium, Kocuria gwangalliensis. The nucleotide sequence of DXS consisted of 2013 bp encoding 671 amino acids residues and turned out to be well conserved during evolution. DXS gene was subcloned into the pColdI expression vector which allows the expression of the recombinant DXS protein with C‐terminal fusion His‐tag and designated to pColdI‐DXS plasmid. The predicted size of recombinant DXS protein is approximately 73 kDa including a C‐terminal His‐Tag fusion. The resulting plasmid was transformed into the E. coli strain BL21(DE3) codon plus cell and the expression of the recombinant DXS protein was induced by the addition of IPTG at a final concentration of 0.1 mM. The expression patterns of recombinant DXS protein were analyzed by 10% SDS‐PAGE.