This study aimed to investigate the effect of X-radiation on the ATPase activity and the activity of antioxidant enzymes, including enzymes of proline biosynthesis, in C. guilliermondii NP-4 yeast cells after X-irradiation and the post-irradiation repair of cells. It has been shown that the X-irradiation of yeasts C. guilliermondii NP-4 at a dose of 300 Gy causes the increase of activity of the enzymes catalase and SOD and the ratio of catalase/SOD. The repair of radiation-induced damages requires a lot of energy, so the demand for ATP increases in irradiated and repaired yeasts. The total and FoF ₁-ATPase activity in yeast homogenates and mitochondria increases after X-irradiation and post-irradiation repair. Amino acid proline is an antioxidative defense molecule and plays a significant role in the adaptation of cells to stress. So higher proline levels in yeasts can protect them from oxidative damage. There are still no systematic studies of the antioxidant properties of proline. In this study, it has been shown that the activity of enzymes of proline biosynthesis (ornithine transaminase and proline-5-carboxylate reductase) in yeasts C.guilliermondii NP-4 increases after X-irradiation and remains higher after post-irradiation repair. Connected with it, the proline amount in X-irradiated and repaired yeasts also is higher than in non-irradiated cells. We suppose that proline has a radioprotective effect on X-irradiated yeasts C. guilliermondii NP-4.