Methionine adenosyltransferase II alpha (MAT2A) is the key enzyme to transform methionine and adenosine-triphosphate (ATP) to S-adenosylmethionine (SAM), a general methyl-group donor in vitro. MAT2A has been reported to participate in the NF-kappa B pathway and maintain the methylated modification, which also affects osteoclastogenesis. In this study, we found the expression of MAT2A was increased upon RANKL stimulation. Pharmacological inhibition of MAT2A by its selective inhibitor AG-270 or genetic silencing by MAT2A-shRNA suppressed osteoclast formation and function in vitro. In vivo treatment with the inhibitor AG-270 also prevented OVX-induced bone loss. Further study revealed that the inhibition of MAT2A affected osteoclast differentiation mainly by suppressing crucial transcription factors and reactive oxygen species induced by RANKL. A quasi-targeted metabolomics assay performed by LC-MS/MS indicated that SAM was reduced by MAT2A knockdown, and the administration of SAM partly rescued the effects of MAT2A inhibition on osteoclastogenesis. These findings revealed that MAT2A is crucial for osteoclastogenesis and might be a potential target for the treatment of osteoporosis attributed to osteoclast dysfunction.