S-adenosylmethionine (AdoMet) synthetase (EC 2.5.1.6) is the enzyme that catalyzes the synthesis of AdoMet, a molecule important for all cellular organisms. We have cloned and characterized an AdoMet synthetase gene (sam1) from Pneumocystis spp. This gene was transcribed primarily as an approximately 1.3-kb mRNA which encodes a protein containing 381 amino acids in P. carinii or P. murina and 382 amino acids in P. jirovecii. sam1 was also transcribed as part of an apparent polycistronic transcript of approximately 5.6 kb, together with a putative chromatin remodeling protein homologous to Saccharomyces cerevisiae, CHD1. Recombinant Sam1, when expressed in Escherichia coli, showed functional enzyme activity. Immunoprecipitation and confocal immunofluorescence analysis using an antipeptide antibody showed that this enzyme is expressed in P. murina. Thus, Pneumocystis, like other organisms, can synthesize its own AdoMet and may not depend on its host for the supply of this important molecule.