MOLECULAR-CLONING AND EXPRESSION OF A GAMMA-INTERFERON-INDUCIBLE ACTIVATOR OF THE MULTICATALYTIC PROTEASE

The multicatalytic protease (MCP) can be activated by two distinct mul tisubunit complexes. One is the regulatory component of the 26 S prote ase, which contains at least 15 distinct subunits. The other is a hexa meric activator composed of 31- and 29-kDa subunits. A cDNA for the sm aller subunit has been cloned and sequenced. The cDNA encodes a protei n of 249 amino acids. Embedded between sequences typical of globular p rotein domains is a stretch of 28 ''alternating'' lysine and glutamic acid residues. Similar regions, which we call KEKE motifs, are also fo und in two MCP subunits, in subunit 12 of the 26 S protease and in a v ariety of chaperonins including hsp90, hsp70, and calnexin. Expression of the activator cDNA in Escherichia coli produced a functional prote in virtually indistinguishable from MCP activator purified directly fr om red blood cells. The recombinant protein formed three isoelectric s pecies on two-dimensional polyacrylamide gel electrophoresis, and it r eacted with antibodies to red blood cell activator. Recombinant activa tor also bound the multicatalytic protease and stimulated cleavage at the carboxyl terminus of hydrophobic or charged residues. Synthesis of the activator subunit was induced by gamma interferon treatment of He La cells. These last two findings have implications for antigen presen tation by class I major histocompatibility receptors.