Improved production of recombinant tumstatin in stably transformed Trichoplusia ni BTI Tn 5B1-4 cells.
We describe the expression and in vitro activity of recombinant tumstatin from stably transformed Trichoplusia ni BTI Tn 5B1-4 cells. Recombinant tumstatin was secreted into a culture medium with a molecular weight of 29 kDa. Recombinant tumstatin was also purified to homogeneity using a simple one-step Ni2+ affinity fractionation. Purified recombinant tumstatin inhibited endothelial cell proliferation in a dose-dependent manner. The concentration at half-maximum inhibition (ED50) for recombinant tumstatin expressed in stably transformed Tn 5B1-4 cells was approximately 0.76 microg/ml. A maximum production level of 4.0 mg/l recombinant tumstatin was obtained in a T-flask culture of Tn 5B1-4 cells, 6 days after cultivation. We also investigated the individual effects of both dimethyl sulfoxide (DMSO) and sodium butyrate on recombinant tumstatin production in stably transformed Tn 5B1-4 cells. Supplementing cultures with DMSO and sodium butyrate separately increased recombinant tumstatin production in stably transformed Tn 5B1-4 cells by 117 and 32%, respectively.
Chang
Kyung Hwa
KH
Department of Genetic Engineering, Kyung Hee University, Suwon 449-701, Republic of Korea.
Lee
Jong Min
JM
Jeon
Hee Kyoung
HK
Chung
In Sik
IS
D000818:Animals; D001324:Autoantigens / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D002087:Butyrates / Q000378:metabolism; D002417:Cattle; D002455:Cell Division / Q000502:physiology; D002478:Cells, Cultured; D024141:Collagen Type IV / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D004121:Dimethyl Sulfoxide / Q000737:chemistry; D042783:Endothelial Cells / Q000166:cytology / Q000378:metabolism; D006801:Humans; D007313:Insecta; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D012997:Solvents / Q000737:chemistry; D014170:Transformation, Genetic
D001324:Autoantigens; D002087:Butyrates; D024141:Collagen Type IV; D011994:Recombinant Proteins; D012997:Solvents; C057112:type IV collagen alpha3 chain; D004121:Dimethyl Sulfoxide