Expression of the core antigen gene of hepatitis B virus (HBV) in Acetobacter methanolicus using broad-host-range vectors

Abstract

Using the broad-host-range promoter probe vector pRS201 for cloning of phage Acm1 promoters, we established a convenient vector system for expression of heterologous genes in different Gram-negative bacteria. The usefulness of this system was demonstrated by expression of the HBV core gene in Acetobacter methanolicus. Plasmids carrying the HBV core gene downstream of different Acm1-phage promoters were transferred to A. methanolicus, a new potential host for recombinant DNA expression. Using enzyme immunoassay and immunoblot techniques, the amount and composition of core antigen produced in A. methanolicus were compared with that derived from Escherichia coli. The expression of immunoreactive core antigen in A. methanolicus exceeds by sevenfold that in E. coli using an expression system with tandemly arranged promoters. Morphological observations by electron microscopy show that the HBV core gene products isolated from both hosts are assembled into regular spherical particles with a diameter of about 28 nm that are comparable to original viral nucleocapsids.

Biochemical characterization and sequence analysis of the gluconate:NADP 5-oxidoreductase gene from Gluconobacter oxydans

“…Expression of plasmid-encoded GNO was reduced but not totally repressed by the intact cI857 repressor in plasmid pRS201P R -gno. In contrast, no functionality of cI857 has been reported for Acetobacter methanolicus (38). The G. oxydans strain harboring pRS201P R -gno had 6.5 times and the G. oxydans strain harboring pRS201P-gno had 85 times the GNO enzyme activity of the wild type.…”

HBV Core Particles as a Carrier for B Cell/T Cell Epitopes

“…Correct folding of the HBc protein and formation of authentic HBc particles have been documented in various mammalian cell cultures [13][14][15][16][17][18], retrovirus [19], vaccinia virus [20,21] and adenovirus [22] expression systems, frog Xenopus oocytes [23], insect Spodoptera cells [24][25][26][27], yeast Saccharomyces cerevisiae [28][29][30][31], in plants Nicotiana tabacum [32], and in bacteria such as Escherichia coli [33][34][35][36][37][38][39][40][41][42], Bacillus subtilis [43], Salmonella [44] and Acetobacter [45]. Electron microscopy revealed the ultrastructural identity of the HBc particles derived from either HBV virions and infected hepatocytes, or from E. coli [46] or yeast [47].…”

On the way toward regulatable expression systems in acetic acid bacteria: target gene expression and use cases

“…In A. methanolica, a RSF1010-based derivative was used to express the core antigen gene of hepatitis B virus subtype ayw under control of the E. coli P lac promoter alone and under tandem control of the strong promoter P acm from the Acetobacter phage Acml and from P lac . When produced from the tandem promoters, the core antigen detected in lysates of A. methanolica was about 7-fold higher than in E. coli (Schröder et al 1991).…”

“…As described before, the AAB genus Acidomonas was established since several characteristics of the former species Acetobacter methanolicus could be distinguished from other type and representative AAB including Acetobacter strains, resulting in the renaming of A. methanolicus to Acidomonas methanolica (Urakami et al 1989). For Acidomonas expression vectors including a basic promoter probe vector have been created based on the popular broad-host-range multicopy plasmid RSF1010 (Schröder et al 1989;Schröder et al 1991). In A. methanolica, a RSF1010-based derivative was used to express the core antigen gene of hepatitis B virus subtype ayw under control of the E. coli P lac promoter alone and under tandem control of the strong promoter P acm from the Acetobacter phage Acml and from P lac .…”