GroES and GroEL are essential chaperones for refolding of recombinant human phospholipid scramblase 1 in E. coli.
Human phospholipid scramblase 1(hPLSCR1), when expressed in E. coli (BL-21 DE3), forms inclusion bodies that are functionally inactive. We studied the effects of various stress inducing agents and chaperones on soluble expression of hPLSCR1 in E. coli (BL-21 DE3). Addition of 3% (v/v) ethanol before induction and decreasing the post-induction temperature to 15 degrees C increased the solubility of hPLSCR1 to approximately 10 and approximately 15% respectively. Presence of groES-groEL complex solubilized the hPLSCR1 to approximately 30% of the total hPLSCR1. Absence of groES-groEL did not improve the solubility of hPLSCR1 suggesting that groES and groEL are the essential chaperones for the correct folding of hPLSCR1 when over-expressed in E. coli.
Sahu
Santosh Kumar
SK
Applied and Industrial Microbiology Laboratory, Department of Biotechnology, Indian Institute of Technology-Madras, Chennai, 600 036, India.
Rajasekharan
Archita
A
Gummadi
Sathyanarayana N
SN
D018835:Chaperonin 10 / Q000378:metabolism*; D018834:Chaperonin 60 / Q000378:metabolism*; D004926:Escherichia coli / Q000187:drug effects / Q000378:metabolism*; D000431:Ethanol / Q000494:pharmacology; D006801:Humans; D049294:Phospholipid Transfer Proteins / Q000737:chemistry* / Q000378:metabolism*; D010743:Phospholipids / Q000378:metabolism; D010957:Plasmids / Q000235:genetics; D017510:Protein Folding / Q000187:drug effects; D020836:Protein Structure, Quaternary; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism*; D012995:Solubility / Q000187:drug effects; D013312:Stress, Physiological / Q000187:drug effects; D013696:Temperature
D018835:Chaperonin 10; D018834:Chaperonin 60; C493422:PLSCR1 protein, human; D049294:Phospholipid Transfer Proteins; D010743:Phospholipids; D011994:Recombinant Proteins; D000431:Ethanol