ABSTRACT Introduction: Although potential relationships with genetic variants of MLCK2, AMPD1 and COL5A1 have been detected in molecular studies evaluating sports performance from the genetic perspective, there are limited data in terms of muscle endurance and physical fitness. Materials and Methods: This study aimed to evaluate these variants in terms of lower limb muscle endurance and physical fitness in thirty-three soccer players. Genotypes were determined by High Resolution Melting (HRM) analysis in qPCR after genomic DNA was isolated from buccal swab samples from the participants. Measurements of lower limb muscle endurance, the dynamic leap and balance test (DLBT), and the standing broad jump test (SBJ) were taken for all the participants. Results: Greater height (p = 0.006), higher DLBT (p = 0.016) and SBJ (p = 0.033) scores, as well as greater left hip adduction (p <0.001), were detected in those with the CT genotype for AMPD1 as compared to those with CC. For MLCK rs28497577, it was found that the players carrying the AA genotype were taller (p = 0.046), heavier (p = 0.049), and had greater left knee extension (p=0.014) and left foot plantar flexion (p =0.040) than those carrying the C allele. Those with the CT genotype for COL5A1 rs12722 had greater right hip extension (p = 0.040) and right knee extension (p = 0.048) than those with the CC genotype. Conclusions: Our results showed that MLCK2 and COL5A1 gene variants are associated with body composition and lower limb muscle endurance, and the presence of the AMPD1 CT genotype may contribute positively to balance, correct positioning, controlled strength, and hip mobility. Evidence level II; Comparative prospective study .