In this study, we aimed to investigate sperm modifications that are related to sperm maturation and capacitation in two shrimp species including the giant freshwater prawn, Macrobrachium rosenbergii, and the black tigher shrimp (Penaeus monodon). In both species, spermatogenesis takes place in the testis and the testicular sperm are then emptied into male reproductive tract, i.e., vas deferens, and the terminal ampoule containing spermatophores. During mating, the spermatophores are deposited into female thelycum which is sperm storage organ in female. We have shown that modification of M. rosenbergii sperm proteins occurred during their storage in male reproductive tract and also in female thelycum. However, sperm protein profiles of sperm taken from the thelycum at 1 h- and 6 h-post insemination were not different, suggesting that modification of sperm proteins in female thelycum may occur rapidly. Moreover, we showed the accumulation of actin in M. rosenbergii sperm in the spike and the base of spike of sperm during maturation process, especially when the sperm were stored in the spermatophore. We also showed changes of tyrosine phosphorylation pattern in sperm during translocation from testis into vas deferens, spermatophores and finally into thelycum of both species. The localization pattern of P. monodon sperm could be used as one of the markers for sperm capacitation in this shrimp species. Proteolytic activities of sperm were also studied. The results showed that P. monodon sperm contained high trypsin-like activities and they gained these activities during storage in female thelycum. The presence of trypsin-like enzymes in sperm of both species was also demonstrated by mean of immunofluorescence staining using soybean trypsin inhibitor. The results suggested the presence of sperm trypsin-like proteases in the subacrosome region of P. monodon and the acrosome region of M. rosenbergii sperm. The sperm protein modifications and proteolytic activities may be important for sperm maturation and capacitation which is the process that make the sperm ready for fertilization.