UBIQUITIN FUSION APPROACH TO HETEROLOGOUS GENE-EXPRESSION IN YEAST - HIGH-LEVEL PRODUCTION OF AMINO-TERMINAL AUTHENTIC PROTEINS

作者：BARR PJ^*; SABIN EA; NG CTL; LANDSBERG KE; STEIMER KS; BATHURST IC; SHUSTER JR

来源：ACS Symposium Series, 1991, 477: 51-64.

The ubiquitin fusion approach to gene expression in eukaryotic systems allows the production of heterologous proteins that are cleaved precisely in vivo from the ubiquitin fusion partner by an endogenous ubiquitin-specific hydrolase. Alternatively, ubiquitin fusions can be isolated from bacterial hosts, and cleaved by the hydrolase in vitro. In each case, the system gives recombinant proteins that contain authentic amino termini. We have found the system to be of particular utility for the production of human gamma interferon (gamma-IFN) and alpha-1-proteinase inhibitor (alpha-1-Pl). We have also used the system for the production of regions of the human immunodeficiency virus type-1 (HlV1) genome that were previously expressed at only low levels in yeast. These include domains of the HlV1 env gene and also the region of the HlV1 pol gene that encodes the HlV1 integrase enzyme. Surprisingly, for one of the env proteins we were able to isolate a product in which the amino-terminal Glu residue was modified by addition of an Arg residue. This arginyl-tRNA-protein-transferase catalyzed process had previously only been observed for short-lived intermediates in the ubiquitin-dependent proteolytic degradation pathway. The HlV1 integrase was found to contain the authentic amino terminus (Phe.Leu.Asn...) previously reported for virion-associated integrase.

• 出版日期1991