Total chemical synthesis of a gene for hepatitis B virus core protein and its functional characterization.
We have chemically synthesized a DNA duplex of 560 nucleotides that codes for the hepatitis B virus (HBV) core protein. The synthetic gene contains 27 unique internal restriction sites. Thereby, it can easily be mutagenized by replacement of rather short restriction fragments. A number of restriction recognition sequences are in common between the synthetic and the authentic gene, thus allowing for the transfer of synthetic segments into the cloned viral genome. Several unexpected mutations in the synthetic gene were readily corrected utilizing the multiple unique restriction sites. In Escherichia coli, the expression level of the synthetic gene product amounts to about 4% of the total soluble protein. It forms particles closely resembling native HBV cores. After transfer of the synthetic gene into the viral genome, transient expression in a hepatoma cell line yields proteins indistinguishable from the native gene products. The synthetic gene thus provides a useful tool for studies on the structure and function of the isolated HBV core protein as well as the gene and its various products in the viral life-cycle.
D001483:Base Sequence; D015153:Blotting, Western; D003001:Cloning, Molecular; D011088:DNA Ligases / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics; D005786:Gene Expression Regulation; D005813:Genes, Synthetic; D005814:Genes, Viral; D005822:Genetic Vectors; D006515:Hepatitis B virus / Q000235:genetics*; D008969:Molecular Sequence Data; D009154:Mutation; D010957:Plasmids; D012150:Polymorphism, Restriction Fragment Length; D014758:Viral Core Proteins / Q000235:genetics*
D014758:Viral Core Proteins; D011088:DNA Ligases