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Characterization of replicative form DNA of the autonomous parvovirus mink enteritis virus
M Shinagawa1, Y Nomura, T Kariatumari
1Department of Veterinary Public Health, School of Veterinary Medicine, Obihiro University of Agriculture.
Abstract:
Characterization of replicative form (RF) DNA of mink enteritis virus (MEV) was carried out. Most of the RF DNA were bound to terminal protein but some were free from the protein. The protein-free RF DNA increased about 7 times from 30 to 50 hr post-infection, while the DNA with protein increased less. The molecules of the replicative intermediate which were partially single-stranded DNA and bound to terminal protein were present. Two terminal conformations, "extended" and "turnaround," were observed in both ends of both terminal protein-bound and protein-free RF DNA. The 5' end labeling revealed that 5' ends of protein-free RF DNA were not blocked to phosphorylation by an amino acid or an oligopeptide which attaches to 5' ends of proteolytically deproteinized RF DNA. Restriction analysis of incomplete RF DNA which was partially double-stranded DNA showed that extended conformation was dominant in such incomplete RF molecules.
Insights
Researchers characterized mink enteritis virus (MEV) replicative form (RF) DNA, finding that protein-free RF DNA increased significantly post-infection. This DNA
Area of Science:
- Virology
- Molecular Biology
- Genetics
Background:
- Mink enteritis virus (MEV) is a parvovirus that causes enteric disease in mink.
- Replicative form (RF) DNA is a double-stranded DNA intermediate crucial for viral replication.
- The role of terminal proteins in MEV RF DNA replication and structure is not fully understood.
Purpose of the Study:
- To characterize the replicative form (RF) DNA of mink enteritis virus (MEV).
- To investigate the association of RF DNA with terminal proteins during infection.
- To analyze the structural conformations and phosphorylation status of MEV RF DNA.
Main Methods:
- Isolation and characterization of MEV RF DNA at various post-infection time points.
- Analysis of protein-bound versus protein-free RF DNA fractions.
- 5' end labeling and restriction enzyme analysis of RF DNA molecules.
- Electron microscopy to observe DNA terminal conformations.
Main Results:
- MEV RF DNA exists in both terminal protein-bound and protein-free forms.
- Protein-free RF DNA significantly increased from 30 to 50 hours post-infection.
- Two distinct terminal conformations ('extended' and 'turnaround') were identified in RF DNA.
- Protein-free RF DNA ends were not blocked for phosphorylation, unlike deproteinized DNA.
Conclusions:
- Terminal proteins play a role in regulating MEV RF DNA replication and structure.
- The increase in protein-free RF DNA suggests a transition phase in the viral replication cycle.
- Understanding MEV RF DNA structure and protein interactions is key to viral replication mechanisms.