Proteasome mutant yeast

The involvement of the 26S proteasome in the degradation of ER proteins was demonstrated by the use of specific inhibitors in mammalian cells (Jensen et al. 1995 Ward et al. 1995) and the investigation of conditional proteasome mutants in yeast (Biederer et al. 1997 Hiller et al. 1996). It is still an open question, how substrate proteins, that have been dislocated from the ER, are targeted to the degrading 26S proteasome. Several observations suggest a localization of 26S proteasomes at the cytosolic side of the ER membrane. Impairing the catalytic activity of the proteasome by specific inhibitors results in the accumulation of ubiquitinated forms of CFTR AF508 in insoluble structures surrounded by ER membranes (Johnston et al. 1998). 

Galan JM, Cantegrit B, Garnier C, Namy O, Haguenauer-Tsapis R (1998) ER degradation of a mutant yeast plasma membrane protein by the ubiquitin-proteasome pathway. FASEB J 12 315-323... 

J. A., Saidowsky, j., Eschee, C., and Wole, D. H. Proteinase yscE, the yeast proteasome/multicatalytic-multifunc-tional proteinase mutants unravel its function in stress induced proteolysis and uncover its necessity for cell survival. EMBO J. 1991, 30, 555-62. 

H., Huber, R., et al. Contribution of proteasomal beta-subunits to the cleavage of peptide substrates analyzed with yeast mutants. J. Biol. Chem. 1998, 273, 25637-25646. 

Cell cycle is one of the physiological processes in which the role of uhiquitin—proteasome-mediated proteolysis is well established. With the advent of yeast mutants that interfered with various phases of the cell cycle, cyclin-dependent kinases (Cdks) were found to have a critical role in regulating the cell cycle. Typically, Cdks activated hy regulatory proteins are known as cyclins. Different Cdk—cyclin complexes are formed at specific stages of the cell cycle such as the S-phase (in which DNA synthesis occurs) and the metaphase. The transition from metaphase to anaphase depends on degradation of cyclins. " " Systematic biochemical studies showed that cyclins were substrates for the uhiquitin—proteasome pathway (Table 5). 

Fig. 2.4 In the p ThrlAla mutant of yeast 20S proteasome, the propeptide is cleaved only at the Arg °-Leu peptide bond. This allowed (using X-ray crystallography) the binding mode of the nonapeptide between the adjacent p and fl subunits to be determined [63].

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