Streptomyces tenjimariensis SS-939, a producer of istamycins, is highly resistant to its own antibiotics and grows in Tryptic Soy Broth containing istamycin A or B at 3, 000 μg/ml. No istamycin-inactivating enzyme was detected in extracts of strain SS-939. Polyphenylalanine synthesis in an in vitro system, consisting of the S-150 fraction and ribosomes prepared from strain SS-939, was not inhibited by 200 μg/ml of istamycins. Using reciprocally reconstituted systems consisting of S-150 fractions and ribosomes from strain SS-939 and those from Streptomyces griseus ISP5236 (istamycin-sensitive strain), ribosomes of strain SS-939 were found to be resistant to istamycins. Thus, ribosomes have the main role in the self-resistance mechanism ofS. tenjimariensis SS-939.