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The structural protein E of the archaeal virus phiCh1: evidence for processing in Natrialba magadii during virus
R Klein1, B Greineder, U Baranyi
1Institute of Microbiology and Genetics, University of Vienna, Dr. Bohr-Gasse 9, Vienna, A-1030, Austria.
Abstract:
phiCh1 is a lysogenic virus for the haloalkalophilic archaeon Natrialba magadii. The virus morphology resembles other members of Myoviridae infecting Halobacterium species. The gene of the major capsid protein E of virus phiCh1 was cloned and the DNA sequence was determined. Gene E was mapped to a 3.2-kbp ClaI fragment, localized to the 5'-end of the phiCh1 genome. The complete nucleotide sequence of this region was determined and the identity of gene E was confirmed by comparing the experimentally determined N-terminal amino acid sequence of the purified protein to the translated DNA sequence of its open reading frame. We present evidence that the gene E product is proteolytically cleaved between Lys(16) and Asn(17) to yield the 305 residue polypeptides found in the mature viral capsid. Processing of the protein itself during virus development was determined by 2D gel electrophoresis using protein E-specific antibodies. Sequence similarity studies revealed an 80% identity to capsid protein Hp32 of phiH, infecting Halobacterium salinarum. RT-PCR analysis as well as Western blot studies revealed gene E as a late gene. Transcripts and proteins could be detected shortly before onset of lysis of the lysogenic strain N. magadii L11.
Insights
The major capsid protein E gene from phiCh1 virus was sequenced and characterized. This study reveals its role as a late gene involved in viral capsid formation in haloalkalophilic archaea.
Area of Science:
- Virology
- Molecular Biology
- Archaea Genetics
Background:
- phiCh1 is a lysogenic virus infecting the haloalkalophilic archaeon Natrialba magadii.
- Its morphology aligns with Myoviridae family viruses that infect Halobacterium species.
Purpose of the Study:
- To clone and determine the DNA sequence of the major capsid protein E gene (gene E) of phiCh1 virus.
- To confirm the identity and processing of the gene E product.
- To investigate the temporal expression of gene E during the viral life cycle.
Main Methods:
- Gene cloning and DNA sequencing
- N-terminal amino acid sequencing of purified protein
- 2D gel electrophoresis with specific antibodies
- Sequence similarity studies
- RT-PCR and Western blot analysis
Main Results:
- Gene E was mapped to the 5'-end of the phiCh1 genome on a 3.2-kbp ClaI fragment.
- The gene E product is proteolytically cleaved to form mature 305-residue capsid polypeptides.
- Gene E exhibits 80% sequence identity to the capsid protein Hp32 of the phiH virus.
- Gene E functions as a late gene, with transcripts and proteins detected before host cell lysis.
Conclusions:
- The study elucidates the genetic and molecular characteristics of phiCh1's major capsid protein E.
- Gene E plays a crucial role in viral capsid assembly and is expressed late in the viral replication cycle.
- This research contributes to understanding virus-host interactions in haloalkalophilic archaea.