Reversed-phase liquid chromatography and sodium dodecyl sulfate polyacrylamide gel electrophoresis characteristics of a recombinant DNA derived malaria antigen.
This paper presents results from analysis of a sample of SK&F 105154 (R32NS1(81], a malaria vaccine candidate produced in Escherichia coli, and discusses some analytical issues of general relevance to the characterization of such products derived from recombinant DNA technology. Anomalous migration and staining behavior were observed in sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Reversed-phase liquid chromatography (RPLC) appeared to resolve four minor components from the principal band, but the minor peaks were found to consist of numerous components resolvable by SDS-PAGE. Western blotting visualized certain components that were not adequately stained by either Coomassie or silver stain. None of the techniques that were employed were individually adequate to characterize the sample, but, taken together, were adequate to characterize the sample and to identify one principal degradation pathway. Degradation within the NS1(81) region decreases the RPLC retention time, while degradation within the R32 segment increases the retention time.
Benedek
K
K
Smith Kline & French Laboratories, King of Prussia, PA 19406-0939.
Hughes
B
B
Seaman
M B
MB
Swadesh
J K
JK
D000595:Amino Acid Sequence; D000818:Animals; D000953:Antigens, Protozoan / Q000032:analysis* / Q000276:immunology; D002851:Chromatography, High Pressure Liquid; D004274:DNA, Recombinant; D004591:Electrophoresis, Polyacrylamide Gel; D008969:Molecular Sequence Data; D010963:Plasmodium falciparum / Q000276:immunology*; D013056:Spectrophotometry, Ultraviolet
D000953:Antigens, Protozoan; D004274:DNA, Recombinant