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Publisher Summary This chapter presents methodology to show that the molecular size heterogeneity of human leukocyte interferon (HLIF) can be eliminated when interferon is produced in the presence of tunicamycin, a specific glycosylation inhibitor. The evidence suggests that the molecular basis of this heterogeneity may be the relative extent of glycosylation of its molecular components. Tunicamycin is used as a specific inhibitor, to probe the glycosylation status of HLIF. Tunicamycin is shown to alter the properties of HLIF molecules that are released into the production media. The use of tunicamycin as a probe, is particularly revealing, because its inhibitory action suggests the N-glycosidic nature of linkage between the polypeptide and its saccharide moiety. HLIF is known to contain two distinct molecular weight components when subjected to sodium dodecyl sulfate-disc gel electrophoresis: 21,000 and 16,000 dalton components in the approximate ratio of 3:7. If HLIF contains sialic acid, then impairment of its glycosylation by tunicamycin should alter its behavior on isoelectric focusing.

Publisher Summary This chapter reveals that human leukocyte interferon (HLIF) is composed of several homologous proteins that are coded for by separate genes in the genome. HLIF can be resolved into two molecular weight components by sodium dodecyl sulfate (SDS)-disc gel electrophoresis and, in addition, has been shown to contain fibroblast-type interferon as a minor component. These three molecular components: (1) 21,000-dalton component (Le21), (2) 16,000-dalton component (Le16), and (3) a fibroblast- type minor component (“F” type) cannot be effectively resolved either by gel filtration or by SDS-disc gel electrophoresis. The chapter describes the efficient separation of these three molecular components, in a single step, by chromatography of HLIF on concanavalin Aagarose. NDV is routinely used as an interferon inducer and human serum is present throughout the production period. It is conceivable that the nature or extent of molecular heterogeneity will vary if these production conditions are changed.