A MATHEMATICAL-MODEL OF N-LINKED GLYCOFORM BIOSYNTHESIS

Metabolic engineering of N-linked oligosaccharide biosynthesis to prod uce novel glycoforms or glycoform distributions of a recombinant glyco protein can potentially lead to an improved therapeutic performance of the glycoprotein product. Effective engineering of this pathway to ma ximize the fractions of beneficial glycoforms within the glycoform pop ulation of a target glycoprotein can be aided by a mathematical model of the N-linked glycosylation process. A mathematical model is present ed here, whose main function is to calculate the expected qualitative trends in the N-linked oligosaccharide distribution resulting from cha nges in the levels of one or more enzymes involved in the network of e nzyme catalyzed reactions that accomplish N-linked oligosaccharide bio synthesis. It consists of mass balances for 33 different oligosacchari de species N-linked to a specified protein that is being transported t hrough the different compartments of the Golgi complex. Values of the model parameters describing Chinese hamster ovary (CHO) cells were est imated from literature information. A basal set of kinetic parameters for the enzyme-catalyzed reactions acting on free oligosaccharide subs trates was also obtained from the literature. The solution of the syst em for this basal set of parameters gave a glycoform distribution cons isting mainly of complex-galactosylated oligosaccharides distributed i n structures with different numbers of antennae in a fashion similar t o that observed for various recombinant proteins produced in CHO cells . Other simulations indicate that changes in the oligosaccharide distr ibution could easily result from alteration in glycoprotein productivi ty within the range currently attainable in industry. The overexpressi on of N-acetylglucosaminyltransferase III in CHO cells was simulated u nder different conditions to test the main function of the model. Thes e simulations allow a comparison of different strategies, such as simu ltaneous overexpression of several enzymes or spatial relocation of en zymes, when trying to optimize a particular glycoform distribution. (C ) 1997 John Wiley & Sons, Inc.