Identification and Preliminary Characterization of a Chitinase Gene in the Autographa californica Nuclear Polyhedrosis Virus Genome

Abstract

A functional chitinase gene (chiA) has been identified in the genome of the Autographa californica nuclear polyhedrosis virus (AcMNPV). It is expressed in the late phase of virus replication in insect cells. High levels of both endo- and exochitinase activity were detected by 12 hr p.i. and remained stable throughout infection. An AcMNPV chiA protein-specific antibody was prepared using recombinant material prepared in bacteria. This was used to demonstrate that a product of approximately 58 kDa was synthesised in virus-infected cells. Immunofluorescence analysis of virus-infected cells showed that most chitinase was located in the cytoplasm. Primer extension analysis of mRNA from AcMNPV-infected cells confirmed that transcription initiated from a baculovirus late start site (TAAG), 14 nucleotides upstream from the putative translation initiation codon. The predicted protein sequence of the AcMNPV chiA shares extensive sequence similarity with chitinases from bacteria and, in particular, the Serratia marcescens chitinase A (60.5% identical residues). Phylogenetic analyses indicate that AcMNPV, or an ancestral baculovirus, acquired the chitinase gene from a bacterium via horizontal gene transfer.

Characterization and phylogenetic analysis of the chitinase gene from the Helicoverpa armigera single nucleocapsid nucleopolyhedrovirus

“…The results indicated that the HaSNPV chitinase is a late gene. Its expression pattern was similar to that of AcMNPV (Hawtin et al, 1995).…”

“…The baculoviral chitinases identified so far are all belonging to family 18 chitinases. It was previously suggested that the AcMNPV chitinase is closely related to the chitinase of S. marcescens (Hawtin et al, 1995). Further phylogenetic analyses, however, could not confirm the proposed lateral transfer from bacteria to baculoviruses (Kang et al, 1998).…”

“…It was extensively studied as a prototype and was found to have endo-and exo-chitinase activities in a wide pH range from 3.0 to 10.0. In AcMNPV-infected Sf21 cells, the expressed chitinase is located in the endoplasmic reticulum (Hawtin et al, 1995(Hawtin et al, ,1997. Chitinases are ubiquitous in nature and have been isolated from bacteria, fungi, nematodes, plants, insects and some vertebrates (Fukamizo, 2000;Koga et al, 1999;Raghaven et al, 1994).…”

Efficient protein production using a Bombyx mori nuclear polyhedrosis virus lacking the cysteine proteinase gene.

“…In addition, the dead body of a CPd-infected larva remains comparatively intact. The chitinase gene in the genome of NPVs (Hawtin et al, 1995) may fulfil a function similar to that of BmNPV-CP.…”

Molecular Cloning and Functional Expression of Chitinase-Encoding cDNA from the Cabbage Moth, Mamestra brassicae

“…Chitinolytic enzymes are essential for chitin turnover and metabolism in chitincontaining organisms, but also exist in higher vertebrates, some plants, bacteria and baculoviruses containing no chitin compounds in relation to self-defense, nutrient assimilation and host invasion (Cohen-Kupiec and Chet, 1998;Collinge et al, 1993;Hawtin et al, 1997;Lyou et al, 2009).…”