Studies on the structures and antigenic properties of rabies virus glycoprotein analogues produced in yeast cells.
We investigated two forms (designated as yGI and yGII) of rabies virus glycoprotein (G) analogues produced in the G cDNA-transfected yeast cells. Molecular weights of yGI and yGII were estimated as 66 and 56 kDa, respectively, according to their relative mobility in SDS-PAGE. Although being produced in large amounts, yGI was present mostly in insoluble forms and hardly extractable with non-ionic detergents. The yGI reacted with polyclonal anti-G antibodies, but did not react with our conformational epitope-specific anti-G monoclonal antibodies (G-MAbs). No protective immunity was induced by yGI in guinea pigs nor in mice. On the other hand, yGII was Triton-soluble, but was only small in amount (at most 1% of total G proteins) and was shown to lack the cytoplasmic domain. The yGII, however, reacted with the G-MAbs and induced protective immunity in guinea pigs as well. When the G-cDNA was expressed in animal cells in culture, a single form (about 66 kDa) of G protein was produced, which displayed similar behaviors as seen in its reactivity with the MAbs and intracellular distribution as seen in the virus-infected cells. These results suggest that most G protein molecules were not processed normally in yeast cells, resulting in abnormal folding and multimer formation, while only a small fraction were occasionally folded normally to have conformational epitopes but were mostly deprived of the C-terminal portion.
Sakamoto
S
S
Research and Development Department, Chemo-Sero-Therapeutic Research Institute, Kumamoto Prefecture, Japan.
Ide
T
T
Tokiyoshi
S
S
Nakao
J
J
Hamada
F
F
Yamamoto
M
M
Grosby
J A
JA
Ni
Y
Y
Kawai
A
A
D000818:Animals; D000956:Antigens, Viral / Q000096:biosynthesis*; D003902:Detergents; D004353:Drug Evaluation, Preclinical; D015967:Gene Expression Regulation, Viral / Q000502:physiology; D005814:Genes, Viral; D005822:Genetic Vectors; D006023:Glycoproteins / Q000276:immunology*; D006031:Glycosylation; D006168:Guinea Pigs; D015151:Immunoblotting; D051379:Mice; D017830:Octoxynol; D011820:Rabies virus / Q000276:immunology*; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D012995:Solubility; D013329:Structure-Activity Relationship