It has been shown that deletion of residue -1 to 7 or 9 residues of rhu IFN -gamma results in the loss of biological activity (H. Hogrefe et al ref. J. Biol. Chem. 21, 12179-86, 1989). Due to the limited amount of material that can be isolated by enzymatic modification, we are using a protein engineering approach to study the relationship between the structure and function of human IFN-gamma. The human IFN-gamma gene has been cloned into PGEM 2 vector and PKK223-3 IFN-gamma gene expressing vector has been constructed. Immunoprecipitation assay and SDS PAGE revealed that cells induced by IPTG produced human IFN-gamma in E. coli JM105. Using PCR and oligonucleotide directed mutagenesis, several mutant genes which contain truncate or replacement at N-terminal will be obtained. Some mutant genes will be cloned into the expression vector, and characterized.
