A dipeptidyl aminopeptidase was identified in JH2SS and was partially purified (approximately 245-fold) by HPLC. Gel filtration chromatography indicated an of 140 000. The partially purified enzyme exhibited a requirement for Co. The pH optimum for the hydrolysis of -Val--Ala--nitroanilide was approximately 9·5. The apparent for this substrate was 0·22 m. The enzyme preferentially hydrolysed X-Ala-Y substrates, but also utilized X-Pro-Y substrates, and therefore is most closely related to the mammalian dipeptidyl aminopeptidase II (EC 3.4.14.-). The enzyme was inhibited by -chloromercuribenzoate, but not by iodoacetate, -ethylmaleimide or the serine protease inhibitor phenylmethylsulphonyl fluoride.