Characterization of a beta-1,3-glucanase encoded by chlorella virus PBCV-1

Sequence analysis of the 330-kb chlorella virus PBCV-1 genome revealed an o pen-reading frame, A94L, that encodes a protein with significant amino acid identity to Glycoside Hydrolase Family 16 beta -1,3-glucanases. The a94l g ene was cloned and the protein was expressed as a GST-A94L fusion protein i n Escherichia coil. The recombinant A94L protein hydrolyzed the beta -1,3-g lucose polymer laminarin and had slightly less hydrolytic activity on beta -1,3-1,4-glucose polymers, lichenan and barley beta -glucan. The recombinan t enzyme had the highest activity at 65 degreesC and pH 8. We predicted tha t the a94l-encoded beta -1,3-glucanase is involved in degrading the host ce ll wall either during virus release and/or is packaged in the virion partic le and involved in virus entry. Therefore, we expected a94l to be expressed late in virus infection. However, contrary to expectations, both the a94l mRNA and the A94L protein appeared 15 min after PBCV-1 infection and disapp eared 60- and 120-min p.i. postinfection, respectively, indicating that a94 l is an early gene. Twenty-seven of 42 chlorella viruses contained the a94l gene. To our knowledge, this is the first report of a virus-encoded beta - 1,3-glucanase. (C) 2000 Academic Press.