One of the fundamental changes during apoptosis is the abnormality of cytoskeleton proteins, which determines some morphological features of apoptotic cells. To understand the role of apoptotic proteases, granzyme B and caspase-3, in the cleavage of the cytoplasmic form of actin, an alternative cell free system based on adult monkey brain tissue was used to reproduce the downstream part of apoptotic program, initiated by the addition of granzyme B. Through extensive Western blot analyses, it was showed that β-actin was cleaved to 41 and 15 ku fragments in the granzyme B-treated brain extract after a 12-hour incubation. The production of these two fragments was further found to be granzyme B dependent. Neither endogenous caspase-3 activated by granzyme B nor its recombinant active form was capable of processing the actin in the brain extract, although the enzyme cleaved the actin purified from rabbit skeleton muscle to produce the 15 ku fragment. The results suggest that endogenous β-actin is resistant to apoptotic proteases, especially to caspase-3, either because of conformational constrains between actin and these proteases or due to the presence of other factors that prevent degradation.