Synthesis, purification and biological properties of a truncated mutant form of human tissue plasminogen activator produced in E. Coli

Abstract

A truncated form of tissue plasminogen activator (t-PA) that lacks the two N-terminal domains of wild type t-PA was efficiently renatured and purified after expression in E. coli. The mutant t-PA was more soluble than wild type t-PA expressed as a nonglycosylated form in E. coli as was predicted by the elimination of a significant segment of predominantly hydrophobic sequence in the mutant. The efficiency of. the renaturation process was, however, unchanged. In vitro biological activity of wild type and mutant t-PA was similar in chromogenic, fibrin plate and clot lysis assays. Both proteins also produced specific plasminogen activation in rabbits, however clearance of the mutant from the circulation was about four times slower than that of wild type t-PA. This confirms previous observations that the N-terminal domains contribute to the rapid clearance of t-PA, but are not essential for specific plasminogen activation in vivo.