Hydrostatic pressure plays an important role in biology. Cells experience high hydrostatic pressure in their native environments through a broad range of mechanical stresses, including stretch, compression, and shear stress. Site-directed spin labeling (SDSL) in combination with electron paramagnetic resonance (EPR) spectroscopy has been developed as an efficient tool to elucidate the structure and the conformational dynamics of macrobiological molecules in various environments. In SDSL, a site with an attached spin label is introduced by site-directed mutagenesis. Typical spin labels are small molecules with an unpaired electron and anisotropic gand A-values, such as nitroxide radicals, trityl-radical radicals, as well as copper, manganese, and gadolinium complexes. Since the EPR spectrum is sensitive to the anisotropic g-and A-values, spin labels based on the nitroxide radicals have been used successfully for probing dynamics and conformational changes of target molecules. SDSL-based EPR spectroscopy has been applied to various biological molecules-including proteins, DNA, and RNA-to probe the local environment, structure, and proximity of individual residues. Recently, the SDSL-based EPR technique has been used to study the stability and heterogeneity of the structural conformation of proteins under high hydrostatic pressure. In particular, McCoy and Hubbell studied the hydrostatic pressure effects on conformational exchange of spin-labeled mutants of T4 lysozyme using the nitroxide-based SDSL and EPR. From the analysis of the nitroxide EPR spectrum, they observed the pressure dependence on the equilibrium states in structural conformations originating from the rotameric exchange. The EPRbased structural analysis requires knowledge of precise values of the g-and A-tensors for the nitroxide spin label. However, experimental determination of these parameters is quite challenging because it is difficult to distinguish spin properties of the spin label itself from the effects of the biological molecule when the spin label is attached to it, and fast tumbling motions of the spin labels makes it difficult to determine them when the study is done with isolated spin labels. Therefore, in practice, the pressure-dependence studies often assume that the g-and A-tensors of the nitroxide radicals are pressure-independent. Accurate quantum-chemistry calculations can fill this gap by providing these essential spinparameters under extreme conditions such as high pressure. In this contribution we use high-level computational methods to characterize the effects of pressure on g-and A-tenors in a nitroxide radical of 2,2,6,6,-tetramethylpiperedine-l-oxyl (TEMPO) molecule. The nitroxide radical is a popular spin-label in EPR experiments. This is the first computational study reporting the effect of hydrostatic pressure on spin properties. Several computational approaches have been developed to study molecular structures under pressure, such as methods employing confining potentials and other means of including pressure in the molecular dynamics and ab initio molecular dynamics simulations. Here, we use eXtended Hydrostatic Compression Force Field (X-HCFF) approach 23 combined with density functional theory (DFT), as implemented in the Q-Chem software package. 24 X-HCFF employs mechanical forces applied perpendicular to the tessellated molecular surface in the process of structure optimization to model the effect of pressure on the molecule. We use X-HCFF to optimize molecular structures under pressures ranging between 0 and 15 GPa. We then use computed structures to calculate g-tensors using coupled-cluster method with single and double excitations (CCSD) combined with response theory and A-tensors using DFT with several functionals. All calculations were carried out with the Q-Chem software package; additional details are given in the supplemental information (SI).