We performed first-principles calculations on the energy, and NMR and polarized infrared (IR) spectra for anhydrous and hydrous aluminous orthoenstatite (OEn) models to help clarify the incorporation mechanisms of Al and OH defects in OEn. Our calculations revealed that proton pairs in M2 vacancies ((2H)M2) adjacent to a tetrahedral Al (AlIV) are energetically more favorable than those remote from Al, and may contribute to the observed correlated 1H NMR peaks near 3.7 and 8.0 ppm, and IR bands near 3550–3570 and 3066 cm−1 (A4 band) for aluminous OEn. Coupled substitutions of AlVI (octahedral Al) + H for 2Mg were found to adopt multiple configurations, and may contribute to the observed IR bands near 3520, 3475 and 3320 cm−1. Coupled substitution of AlIV + H for 1Si may contribute to the observed IR band near 3380–3400 cm−1. 4H in SiB vacancies ((4H)SiB) adjacent to an AlVI were found to be energetically more favorable than those remote from Al, and may be the origin for an IR band observed near 3600–3620 cm−1. These results allow the incorporation mechanisms of water in synthetic and natural aluminous orthopyroxenes to be deciphered from the available NMR and IR data, and suggest that both (2H)M2 defects associated with Al and simultaneous coupled substitutions of Al + H for 2Mg and 1Si contribute to the observed correlation between Al and water incorporation, and the nearly unity AlIV/AlVI ratio. (4H)SiB defects associated with Al may also be present in some synthetic OEn and mantle-derived orthopyroxene.