Spermatogenesis is a dynamic process that culminates in the production of mature spermatozoa in the seminiferous tubules of sexually mature animals. Although sperm leaving the testis are fully differentiated, they must further undergo two additional maturation steps before acquiring the capability to fertilize the egg. Such processes take place during the epididymal residency and transport in the seminal fluid during ejaculation and, after delivery into the female reproductive tract, during the journey aiming the encountering the egg in the oviduct. Throughout this trip, spermatozoa are exposed to different reproductive fluids whose molecular compositions regulate the progress towards obtaining a fertilized competent cell. This review summarizes the evidence obtained so far supporting the participation of male and female reproductive tract-derived proteins in the modulation of sperm fertilizing ability and discusses the mechanisms by which such regulation may be accomplished.