Rapid degradation of solid‐phase bound peptides by the 20S proteasome

Proteasomes are cellular proteases involved in the degradation of numerous cellular proteins. The 20S proteasome is a cylindrical 28‐mer protein complex composed of two outer heptameric α‐rings forming the entrance for the protein substrate and two inner heptameric β‐rings carrying the catalytic sites. Numerous in vitro studies have provided evidence that the 20S proteasome may degrade peptides of various lengths and even unfolded full‐length polypeptide chains. However, a direct demonstration that the 20S proteasome may also cleave surface‐attached immobilized peptides is lacking so far. To this end, we used a model system by coupling peptides from different source proteins covalently to the surface of glass beads and applied nanoLC/MS analysis to monitor the generation of proteolytic fragments in the presence of the 20S proteasome. Detectable amounts of cleavage products occurred within a few minutes indicating a much higher cleavage rate than observed with the same substrates in solution. Our finding lends support to the idea that proteasomes may directly degrade segments of membrane‐bound proteins protruding into the aqueous phase. Copyright © 2013 European Peptide Society and John Wiley & Sons, Ltd.