Efficient production and isolation of recombinant amino-terminal half-molecule of human serum transferrin from baby hamster kidney cells.
Expression of the amino-terminal lobe of human serum transferrin secreted into the culture medium by transformed baby hamster kidney (BHK) cells has been increased from the levels reported originally of 10-15 micrograms/ml to 55-120 micrograms/ml. Use of the serum substitute, Ultraser G, has facilitated isolation of the recombinant protein, resulting in approximately 80% recovery of expressed hTF/2N from the culture medium. In the three experiments described, 300-750 mg of recombinant protein was collected over a period of 25 days from five expanded surface roller bottles each containing 200 ml of medium (seven to nine collections). The use of alginate beads to encapsulate the transformed BHK cells provided no advantage over normal culturing over 25 days. A lag in production resulting in 30% less recombinant protein over this time period was observed. The production and isolation procedures described are easily handled by one person. The system is amenable to incorporation of isotopically substituted amino acids useful in NMR studies.
Mason
A B
AB
Department of Biochemistry, University of Vermont College of Medicine, Burlington 05405.
Funk
W D
WD
MacGillivray
R T
RT
Woodworth
R C
RC
D000818:Animals; D002478:Cells, Cultured; D002848:Chromatography, DEAE-Cellulose; D006224:Cricetinae; D003636:DEAE-Cellulose / Q000031:analogs & derivatives; D015870:Gene Expression; D006801:Humans; D010957:Plasmids; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification; D014168:Transferrin / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification; D014170:Transformation, Genetic
D011994:Recombinant Proteins; D014168:Transferrin; D003636:DEAE-Cellulose