PROLINE-137 IS CRITICAL FOR ADENOVIRUS PROTEASE ENCAPSIDATION AND ACTIVATION BUT NOT ENZYME-ACTIVITY

Infection with the adenovirus type 2 ts1 mutant at the nonpermissive t emperature resulted in the production of noninfectious virions. This h as been ascribed to the P137L mutation in the virus-encoded cysteine p rotease which causes a defect in protease activity. Here we have exami ned the ts1 defect in detail as a means of learning more about the vir al protease. The ts1 protease accumulated in the nucleus normally and was found associated with incomplete particles as was the case with wt . This enzyme was active in both wt and ts1 incomplete particles produ ced at 39 degrees (ts1-39 TCs), provided they were dissociated with 4 M urea. While the wt protease was packaged into complete particles, th e ts1-39 particles were totally devoid of protease. This defect was ne arly completely corrected by addition of the 11-residue activating pep tide PVIc (GVQSLKRRRCF) to the medium late in infection. Rescue of ts1 occurred via restoration of enzyme activity and packaging of the ts1 enzyme into complete virions. Recombinant ts1 enzyme was not temperatu re sensitive. The P137L mutation responsible for the ts1 defect appear ed therefore to be an in vivo phenotype involving apparently linked ev ents of protease packaging and activation mediated by the PVI protein. (C) 1995 Academic Press, Inc.