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Phosphorylation of avian retrovirus matrix protein by Ca2+/phospholipid-dependent protein kinase
J Leis1, N Phillips, X Fu
1Department of Biochemistry, Case Western Reserve University School of Medicine, Cleveland, OH 44106.
Abstract:
The matrix protein from avian myeloblastosis virus and the Rous sarcoma virus, Prague C strain, is a phosphoprotein. A comparison of the amino acid sequences shows these phosphoproteins are very similar. The sites of phosphorylation of the matrix protein purified from virions are identified as serine residues 68 and 106. Treatment with purified rabbit skeletal-muscle protein phosphatase 1 or 2A, selectively releases phosphate from serine 68, while alkali treatment releases phosphate from both sites. When analyzed as a substrate for six different protein kinases, only the Ca2+/phospholipid-dependent protein kinase modifies the matrix protein. The serine residues phosphorylated in vivo are identical to those phosphorylated in vitro by this protein kinase. The role of these phosphorylation events in viral production is discussed.
Insights
The matrix phosphoprotein from avian myeloblastosis virus and Rous sarcoma virus is highly similar. Specific serine residues are phosphorylated by a Ca2+/phospholipid-dependent protein kinase, impacting viral production.
Area of Science:
- Virology
- Molecular Biology
- Protein Chemistry
Background:
- The matrix protein of avian myeloblastosis virus (AMV) and Rous sarcoma virus (RSV) Prague C strain are phosphoproteins.
- Amino acid sequence comparison reveals significant similarity between these viral matrix phosphoproteins.
Purpose of the Study:
- To identify the specific sites of phosphorylation on the matrix phosphoprotein.
- To investigate the kinases responsible for phosphorylating the matrix protein in vitro and in vivo.
- To understand the role of these phosphorylation events in viral production.
Main Methods:
- Purification of matrix phosphoprotein from virions.
- Amino acid sequencing and analysis.
- Phosphatase treatment (protein phosphatase 1 and 2A) to identify phosphorylation sites.
- In vitro kinase assays using various protein kinases, including Ca2+/phospholipid-dependent protein kinase.
- Comparison of in vitro and in vivo phosphorylation sites.
Main Results:
- Serine residues 68 and 106 were identified as the primary sites of phosphorylation on the matrix protein.
- Protein phosphatase 1 and 2A selectively removed phosphate from serine 68.
- Alkali treatment released phosphate from both serine 68 and 106.
- Only Ca2+/phospholipid-dependent protein kinase phosphorylated the matrix protein in vitro.
- In vivo and in vitro phosphorylation sites were identical.
Conclusions:
- The matrix phosphoprotein's phosphorylation sites are precisely regulated.
- Ca2+/phospholipid-dependent protein kinase is likely responsible for in vivo phosphorylation of the matrix protein.
- These phosphorylation events are crucial for viral production, suggesting a regulatory role.