Figure 6.

Media Supplementation Can Potentially Bypass the NSD Biosynthetic Pathway to Partially Mitigate Galactosylation and Sialylation Bottlenecks

(A) Supplementation of manganese to enhance B4galt1 activity alone is insufficient to reduce the bottleneck at galactosylation level, whereas galactose supplementation (5 g/L) leads to 25- to 50-fold increase in intracellular pool of UDP-Gal (left), resulting in sustained levels of galactosylation throughout the cell culture (right).

(B) Supplementation of 20 mM ManNAc bypasses the self-regulation on CMP-Sia biosynthesis (shown by dotted arrow) and leads to ∼30-fold increase in intracellular concentrations of CMP-sialic acid (left), resulting in around 2-fold increase in sialylated species.

(C) Temporal variations in the intracellular concentrations of the NSD substrates and/or potential substrate competition for intra-Golgi transport due to shared transport protein might result in the temporal heterogeneity in glycoforms observed in fed-batch cultures.