[Cloning, expression, and isolation from Escherichia coli of human protein SURF-6 translationally fused to glutathione S-transferase].
cDNA of human gene Surf-6 (hSutf-6) was amplified and cloned into vector pGEX-2T for the expression in the bacterial system of protein hSURF-6 translationally fused to glutathione S-transferase. The resulting vector is named as pGEX-2T-GST-hSurf-6. Superproducer of chimeric protein GST-hSURF-6 was obtained on the basis of Escherichia coli strain BL21-CodonPlus(DE3)-RIL. Its purification was performed by the affinity chromatography on L-glatathione-sepharose. The proportion of recombinant protein GST-hSURF-6 in the optimized conditions was not less than 15% of the total bacterial protein, and up to 7 mg of the protein was isolated from 1 liter of culture of the producer strain. The final fraction of eluate contained approximately 80% of GST-hSURF-6. The amount and the purity of the isolated protein were sufficient to immunize animals and obtain antibodies. Protein GST-hSURF-6 can also be used as an affinity ligand for revealing protein partners of hSURF-6 in human cells.
M Iu
MIu
Kordiukova
O V
OV
Zatsepina
M A
MA
Polzikov
D000917:Antibody Formation; D015153:Blotting, Western; D002846:Chromatography, Affinity; D002847:Chromatography, Agarose; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics*; D005822:Genetic Vectors; D005982:Glutathione Transferase / Q000235:genetics; D006367:HeLa Cells; D006801:Humans; D009687:Nuclear Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification; D017422:Sequence Analysis, DNA
D009687:Nuclear Proteins; D011993:Recombinant Fusion Proteins; C484378:SURF6 protein, human; D005982:Glutathione Transferase