MOLECULAR-CLONING AND EXPRESSION OF SUBUNIT-9 OF THE 26S PROTEASOME

Seven peptides from subunit 9 (S9) of the human 26S proteasome were se quenced and this information was used to clone a HeLa cDNA that encode s the 46 kDa subunit. Rabbit polyclonal antisera were made against a u biquitin fusion protein containing 12 amino acids from S9 and against a full-length S9 expressed in E. coli. Western blot analysis showed th at the S9-specific antibodies bound the 26S proteasome and its regulat ory complex separated on non-denaturing gels. In SDS-PAGE samples of t he two complexes, the S9-specific antibodies bound a single 46 kDa sub unit. Thus, a cDNA encoding a novel 26S protease subunit has been isol ated, sequenced, and expressed. (C) 1997 Federation of European Bioche mical Societies.