The steric effects imposed by the isopropyl group of valine in the conformational stabilization of the capped dipeptide N-acetyl-L-valinamide (Ac-Val-NH₂) have been studied by laser ablation molecular beam Fourier transform microwave (LA-MB-FTMW) spectroscopy. The rotational and quadrupole coupling constants of the two ¹⁴N nuclei determined in this work show that this dipeptide exists as a mixture of C₇ and C₅ conformers in the supersonic expansion. The conformers are stabilized by a CO⋯H–N intramolecular hydrogen bond closing a seven- or a five-membered ring, respectively. The observation of both conformers is in good agreement with previous results on the related dipeptides containing different residues, confirming that the polarity/non-polarity of the side chains of the amino acid is responsible for the conformational locking/unlocking. The voluminous isopropyl group is not able to prevent the less stable C₅ conformer from forming but it destabilizes the CO⋯H–N interaction.