EFFICIENT EXPRESSION AND RAPID PURIFICATION OF HUMAN T-CELL LEUKEMIA-VIRUS TYPE-1 PROTEASE

Human T-cell leukemia virus type 1 (HTLV-1) is an oncovirus that is cl inically associated with adult T-cell leukemia. We report here the con struction of a pET19-based expression clone containing HTLV-1 protease fused to a decahistidine-containing leader peptide. The recombinant p rotein is efficiently expressed in Escherichia coli, and the fusion pr otein can be easily purified by affinity chromatography. Active mature protease in yields in excess of 3 mg/liter of culture can then be obt ained by a novel two-step refolding and autoprocessing procedure. The purified enzyme exhibited K-m and K-cat values of 0.3 mM and 0.143 sec (-1) at pH 5.3 and was inhibited by pepstatin A.