R. Gueckel et al., MUTATIONS IN THE YEAST PROTEASOME BETA-TYPE SUBUNIT PRE3 UNCOVER POSITION-DEPENDENT EFFECTS ON PROTEASOMAL PEPTIDASE ACTIVITY AND IN-VIVO FUNCTION, The Journal of biological chemistry, 273(31), 1998, pp. 19443-19452
Proteasomes are highly complex proteases responsible for selective pro
tein degradation in the eukaryotic cell. 26 S proteasomes consist of t
wo regulatory 19 S cap complexes and the 20 S proteasome, which acts a
s the proteolytic core module. We isolated six mutants of the yeast Sa
ccharomyces cerevisiae containing mutations in the 20 S proteasome P-t
ype subunit Pre3. Three mutations (pre3-2, pre3-3, and pre3-5) which r
eside at the active site cleft of the Pre3 subunit solely caused reduc
tion of the proteasomal peptidylglutamyl peptide-hydrolyzing activity
but did not lead to detectable defects in protein degradation nor to a
ny other phenotype. However, the pre3-2 mutation strengthened phenotyp
es induced by other 20 S proteasomal mutations, indicating that the pe
ptidylglutamyl peptide-hydrolyzing activity has to fulfill some rescue
functions. The other three mutations (pre3-1,pre3-4 and pre3-6) are l
ocated at diverse sites of the Pre3 protein and caused multiple defect
s in proteasomal peptide cleaving activities. pre3-1 and pre3-6 mutant
s exhibited significant defects in proteasomal protein degradation; th
ey accumulated ubiquitinated proteins and stabilized defined substrate
proteins as, e.g. fructose-1,6-bisphosphatase. In addition, pre3-1 an
d pre3-6 mutant cells exhibited pleiotropic phenotypes as temperature
sensitivity and cell cycle-related effects.