| Abstract: | Expression of the coat protein gene of RNA bacteriophage fr in Escherichia coli cells leads to the formation of capsid-like structures of ca. 25 nm in diameter, which are immunologically indistinguishable from the native phage fr capsids. The modification strategy of the coat protein gene by gene engineering technique was developed in order to localize coat protein regions, which are exposed on the capsid surface and are capable to include foreign amino acid inserts without an appreciable effect on the capsid self-assembly. The oligonucleotide linkers, coding short amino acid sequences and bearing also convenient restriction sites, were synthesized and inserted into different regions of the coat protein gene. The mutant proteins, containing insertions of 2-12 amino acids in potentially exposed regions, were obtained. It was shown that N- and C-terminal insertions, as well as the insertion into codon 51 in the RNA-binding region, do not prevent the self-assembly. The regions (codons 96 and 112) were also revealed, insertions in them decreased drastically the protein yield as a consequence of a block in the self-assembly. |
