Production of a membrane-bound protein, the human gamma-glutamyl transferase, by CHO cells cultivated on microcarriers, in aggregates and in suspension.
Recombinant Chinese Hamster Ovary (CHO) cells, engineered for the production of human gamma-glutamyl transferase (GGT), have been grown on Cytodex 1 microcarriers, as aggregates, or as single cells in suspension after adaptation. GGT is a membrane bound enzyme which was not secreted during the culture period. The maximal enzyme activity was found to be directly related to the achieved maximal cell density. Culture of CHO on microcarriers yielded the fastest growth, with a specific growth rate of 0.04 h-1, the highest cell density (near 1.3 x 10(6) cells ml-1), and the highest enzyme activity around 300 mU ml-1, which corresponded to a specific cellular level of 20 mU 10(-5) cells. GGT could also be produced by growing CHO cells in suspension as single cells or as aggregates. Under these conditions, however, the specific CHO growth rate was significantly slower and the GGT level per cell was divided by a factor 6. Growing CHO cells without microcarriers also resulted in differences in cell metabolism, with a higher conversion yield of glutamine into ammonia, and a higher cell lysis. The catalytic kinetic constants of the enzyme were found identical for the three culture systems.
Chevalot
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I
Laboratoire des Sciences du Génie Chimique, UPR CNRS 6811, Nancy.
Visvikis
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A
Nabet
P
P
Engasser
J M
JM
Marc
A
A
D000222:Adaptation, Physiological; D000818:Animals; D016466:CHO Cells; D002449:Cell Aggregation / Q000502:physiology; D002455:Cell Division / Q000502:physiology; D006224:Cricetinae; D003911:Dextrans; D006801:Humans; D007700:Kinetics; D008565:Membrane Proteins / Q000096:biosynthesis*; D008863:Microspheres; D015202:Protein Engineering; D005723:gamma-Glutamyltransferase / Q000096:biosynthesis*
D003911:Dextrans; D008565:Membrane Proteins; C052839:Cytodex; D005723:gamma-Glutamyltransferase