Contaminating fungi, such as Fusarium species, produce metabolites that may interfere with normal barley grain proteolysis pattern and consequently, affect malt and beer quality. Protein compositional changes of an initial mixture of 20 % Fusarium culmorum infected and 80 % noninfected mature barley grains and respective malt are reported here. Proteolytic activity of infected barley grains (IBG) and respective malt, with controls (uninfected grains), were characterized using protease inhibitors from each class of this enzyme, including metallo-, cysteine, serine, and aspartic proteases, as well as uninhibited protease fractions. The proteins were extracted according to the Osborne fractionation and separated by size exclusion chromatography. Additionally, two-dimensional (2D) gel electrophoresis (GE) was used to analyze hydrophobic storage proteins isolated from the control and IBG. Analyses revealed that F. culmorum IBG had a twofold increase of proteolytic activity compared to the control sample, which showed an increase in all protease classes with aspartic proteases dominating. Infected and control malt grains were comparable with cysteine proteases representing almost 50 % of all proteolytic enzymes detected. Protein extractability was 31 % higher in IBG compared to the control barley. The albumin fraction showed that several metabolic proteins decreased and increased at different rates during infection and malting, thus showing a complex F. culmorum infection interdependence. Prolamin storage proteins were more hydrophobic during barley fungal infection. F. culmorum interfered with the grain hydrolytic protein profile, thereby altering the grain's protein content and quality.