Doctoral Dissertations

Date of Award

3-1981

Degree Type

Dissertation

Degree Name

Doctor of Philosophy

Major

Microbiology

Major Professor

David A. Brian

Committee Members

R. Courtney, S. Kennel, A. Brown

Abstract

The tissue culture-adapted strain (Mebus) of bovine coronavirus was grown in the presence of isotopically-labeled amino acids, glucosamine, or orthophosphate, for the purpose of analyzing the virion structural polypeptides. Five species of polypeptides were identified when purified virions were solubilized in urea and sodium dodecyl sulfate and resolved by polyacrylamide gel electrophoresis. Four species were labeled with 3[H]-glucosamine and are designated gpl40 (molecular weight = 140,000), gpl20 (molecular weight = 120,000), gp100 (molecular weight = 100,000), and gp25 (molecular weight = 26,000). Pp52 (molecular weight = 52,000) was labeled with 32[P]- orthophosphate. Gpl40, pp52 and gp26 were the most abundant species. Gpl40 is apparently a polymer of 65,000 molecular weight glycopolypeptides (designated gp65) held together by disulfide linkages. All four glycoproteins were susceptible to proteolytic cleavage and enzymatic iodination when intact virions were studied and are thus at least partially exposed on the virion surface. Pp52, in the intact virion, was unaffected by protease and was not enzymatically iodinated. It is, therefore, presumed to be the internal RNA-associated protein.

Two lines of evidence suggested that gpl40 is the virion hemagglutinin protein. (1) Purified virions digested with bromelain lost gpl20, gp100, and gp26, but retained both gpl40 (observed as gp65 on reducing gels) and the internal pp52 along with full hemagglutinating activity. (2) Purified virions treated with NP40 became subviral particles that retained pp52 and gp26, but no hemagglutinating activity. Hemagglutination was found in the NP-40 soluble fraction along with gpl40, gpl20, and gp100. The only intact molecule that was common in both circumstances of hemagglutination was the gpl40 polypeptide.

Download
Files over 3MB may be slow to open. For best results, right-click and select "save as..."

Share

COinS