Proteasome is a multicatalytic enzyme complex responsible for proteolysis of damaged proteins and an important target for drug discovery in the pharmaceutical industry. Development of fast and economic strategies for detection of proteasome activity and inhibition is a topic of intensive research. The activity of the 20S proteasome was investigated by voltammetry and atomic force microscopy. The hydrolysis of peptide bonds was studied in incubated solutions of proteasome with oligopeptide sequences specific to each chymotrypsin, trypsin and caspase activity of proteasome, before and after inhibition with epoxomicin. The time-dependence of the proteolysis and the effect of substrate and inhibitor concentrations on the rate of enzymatic reaction were investigated. Different interaction mechanisms were characterized and enzyme kinetic parameters determined. The adsorption patterns of reaction mixture components were characterized by atomic force microscopy in order to understand the processes when saturation of enzyme catalytic centres occurs for high substrate concentrations. (C) 2017 Elsevier B.V. All rights reserved.