The broadband rotational spectrum of methyl butyrate from 8 to 18 GHz, recorded using a chirp-pulsed Fourier transform microwave (FTMW) spectrometer, was combined with high resolution FTMW measurements over the 2–26.5 GHz region to provide a comprehensive account of its microwave spectrum under jet-cooled conditions. Two low-energy conformers, one with a fully extended, heavy-atom planar anti/anti structure (a, a), and the other with a gauche propyl chain (g±, a), were assigned in the spectrum. Torsional A/E splittings due to the internal rotation of the methoxy methyl group were resolved for both lower energy conformers, and were fitted using the program XIAM and BELGI, providing an estimate of the barrier to methyl internal rotation of V3 ≈ 420 cm−1. The conformational landscape of methyl butyrate occurs on a two-dimensional potential energy surface, which was mapped out by quantum chemical calculations at the B2PLYP-D3BJ/aug-cc-pVTZ level of theory. The low torsional barrier about the C-C(=O)O bond leads to collisional removal of population originally in the (a, g±) and (g±, g±) minima into the (a, a) and (g±, a) minima, respectively, during the cooling in the expansion. Analysis of experimental intensities in the spectrum provide percent populations downstream in the expansion of 41 ± 4% (a, a), and 59 ± 6% (g±, a).