The ACTN3 (R577X) gene encodes for a structural protein that is exclusively expressed in the Z-disc of type II muscle fibers. Homozygosis (577XX) for the stop codon in the ACTN3 polymorphism results in alpha-actinin-3 complete deficiency. Previous studies have shown low frequencies of the ACTN3 XX genotype in elite sprinters compared to the general population. This study tests 35 Italian elite gymnasts and 53 controls. ACTN3 XX genotype (2.8% vs. 18.8%; p < 0.04) and X allele (27.1% vs. 43.3%; p < 0.04) frequencies were significantly lower in gymnasts compared to controls. The ACTN3 XX genotype was underrepresented in female and male gymnasts compared to controls, but was only significant for males (male: 0% vs. 16.1%, p < 0.04; female: 5.5% vs. 22.7%, p = 0.39). These results suggest that alpha-actinin-3 is beneficial to skeletal muscle function in generating forceful contractions at high velocity. In conclusion, our results associated the ACTN3 R577X polymorphism with male and possibly female elite gymnastic performance.