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Deduced sequence of the bovine coronavirus spike protein and identification of the internal proteolytic cleavage site
S Abraham1, T E Kienzle, W Lapps
1Department of Microbiology, University of Tennessee, Knoxville 37996-0845.
Abstract:
The sequence of the spike (also called peplomer or E2) protein gene of the Mebus strain of bovine coronavirus (BCV) was obtained from cDNA clones of genomic RNA. The gene sequence predicts a 150,825 mol wt apoprotein of 1363 amino acids having an N-terminal hydrophobic signal sequence of 17 amino acids, 19 potential N-linked glycosylation sites, a hydrophobic anchor sequence of approximately 17 amino acids near the C terminus, and a hydrophilic cysteine-rich C terminus of 35 amino acids. An internal Lys-Arg-Arg-Ser-Arg-Arg sequence predicts a protease cleavage site between amino acids 768 and 769 that would separate the S apoprotein into S1 and S2 segments of 85690 and 65153 mol wt, respectively. Amino terminal amino acid sequencing of the virion-derived gp 100 spike subunit confirmed the location of the predicted cleavage site, and established that gp 120 and gp 100 are the glycosylated virion forms of the S1 and S2 subunits, respectively. Sequence comparisons between BCV and the antigenically related mouse hepatitis coronavirus revealed more sequence divergence in the putative knob region of the spike protein (S1) than in the stem region (S2).
Insights
Researchers sequenced the bovine coronavirus (BCV) spike protein gene. This revealed its structure, including cleavage sites and glycosylation, and differences compared to mouse hepatitis coronavirus.
Area of Science:
- Virology
- Molecular Biology
- Protein Chemistry
Background:
- Bovine coronavirus (BCV) is an important animal pathogen.
- The spike (S) protein mediates viral entry and is a key determinant of tropism and antigenicity.
- Understanding the BCV S protein structure is crucial for developing diagnostics and therapeutics.
Purpose of the Study:
- To determine the complete gene sequence of the Mebus strain of BCV spike protein.
- To analyze the predicted protein structure, including glycosylation sites and cleavage points.
- To compare the BCV S protein sequence with that of a related coronavirus.
Main Methods:
- cDNA cloning of genomic RNA from the Mebus strain of BCV.
- Gene sequencing and bioinformatic analysis to predict protein structure.
- Amino terminal amino acid sequencing of virion-derived spike subunits.
- Sequence comparison with mouse hepatitis coronavirus (MHV).
Main Results:
- The BCV S apoprotein consists of 1363 amino acids with predicted signal and anchor sequences, and 19 N-linked glycosylation sites.
- A protease cleavage site was identified, separating the S apoprotein into S1 and S2 subunits (gp120 and gp100, respectively).
- Sequence comparison revealed greater divergence in the S1 (knob) region than in the S2 (stem) region between BCV and MHV.
Conclusions:
- The deduced sequence and structural features of the BCV spike protein provide insights into its function and evolution.
- The identified cleavage site and glycosylation patterns are important for understanding BCV assembly and infectivity.
- Comparative sequence analysis highlights distinct evolutionary trajectories of the spike protein in different coronaviruses.