Glutamine-based dipeptides are utilized in mammalian cell culture by extracellular hydrolysis catalyzed by a specific peptidase.
A murine hybridoma (CC9C10) was grown in media containing alanyl-glutamine (ala-gln) or glycyl-glutamine (gly-gln) as a substitute for glutamine. High cell yields were obtained in the presence of 6 mM ala-gln or 20 mM gly-gln. The final cell yield in gly-gln was 14% higher than in gln. Monoclonal antibody productivity was comparable in gln, ala-gln or gly-gln. Substrate utilisation and metabolism was affected by the presence of the dipeptides, particularly with gly-gln. The specific consumption rates of glucose and six amino acids were reduced. Also the accumulation of ammonia and lactate was significantly lower. The higher concentration of gly-gln was necessary for cell growth because of the presence of a peptidase with a lower affinity for gly-gln. The peptidase activity of the basal medium and serum was low. However, a substantial peptidase activity was found in the cytosolic fraction of the cells with an apparent Km of 1.2 mM with respect to ala-gln and 14 mM with respect to gly-gln. It is proposed that the mechanism of dipeptide utilisation involves extracellular hydrolysis by this peptidase following its release into the culture medium.
Christie
A
A
Department of Microbiology, University of Manitoba, Winnipeg, Canada.
Butler
M
M
D000596:Amino Acids / Q000032:analysis; D000641:Ammonia / Q000378:metabolism; D000818:Animals; D002384:Catalysis; D002455:Cell Division; D017077:Culture Media, Conditioned; D004151:Dipeptides / Q000737:chemistry / Q000378:metabolism*; D005947:Glucose / Q000378:metabolism; D005973:Glutamine / Q000378:metabolism*; D006825:Hybridomas; D006868:Hydrolysis; D007700:Kinetics; D007773:Lactates / Q000378:metabolism; D019344:Lactic Acid; D051379:Mice; D010447:Peptide Hydrolases / Q000378:metabolism*; D013242:Sterilization; D013379:Substrate Specificity
D000596:Amino Acids; D017077:Culture Media, Conditioned; D004151:Dipeptides; D007773:Lactates; D005973:Glutamine; D019344:Lactic Acid; D000641:Ammonia; D010447:Peptide Hydrolases; D005947:Glucose