Walter Mitsuo Nakatsukasa

A83543A-D, unique fermentation-derived tetracyclic macrolides

Abstract A multi-factored complex of structurally-unique macrolides was isolated from culture broths of a new species of Saccharopolyspora. The core structure consists of a 5,6,5-cis-anti-trans-tricyclic ring system fused to a 12-membered macrocyclic lactone, which is further substituted by an amino- and a neutral sugar.

Development of cloning vehicles from the Streptomyces plasmid pFJ103.

A 20-kb plasmid, pFJ103, was isolated from a strain of Streptomyces granuloruber. A restriction endonuclease map of the plasmid was constructed. A Streptomyces gene that specifies resistance to the antibiotic thiostrepton was subcloned into Escherichia coli plasmid pBR322, inserted into pFJ103 and transformed into Streptomyces ambofaciens protoplasts. Two classes of transformants were obtained. One carries the pFJ104 plasmid consisting of the entire pFJ103 with the 1.8-kb thiostrepton resistance gene insert. The other carries the pFJ105 plasmid consisting of the 2.9-kb replicon segment of pFJ103 with the same thiostrepton resistance insert. A gene for neomycin resistance together with the entire E. coli pBR322 plasmid were cloned into pFJ105. The resulting E. coli-Streptomyces bifunctional vector, pFJ123, transformed both E. coli and Streptomyces. The small size of pFJ105, its ease of isolation, and efficient transformation of Streptomyces protoplasts establishes it, and its derivatives, as useful plasmid cloning vehicles for fundamental and applied studies.

A82775b and a82775c, novel metabolites of an unknown fungus of the order sphaeropsidales

Compounds A82775B (1) and A82775C (2) were isolated as non-antimicrobial by-products during attempts to isolate an elusive G+, G- antibiotic from culture A82775. an unidentified mold. A82775 factors B and C are present in both the mycelium and filtered broth. The metabolites were recovered from the filtered broth by adsorption on Diaion HP-20 nonfunctionalized macroreticular resin and purified by preparative reverse phase HPLC on C18 bonded phase supports. The structure elucidation of A82775 factors B (1) and C (2) has been accomplished by spectroscopic means including COSY, NOESY, INAPT, HMQC and 2D-INADEQUATE NMR experiments.

pFJ265, a new cloning vehicle for Streptomyces

A 9.3-kb plasmid, pNM100, was isolated from Streptomyces virginiae (NRRL 15156) and characterized. Streptomyces genes for thiostrepton and neomycin resistance were cloned into pNM100 to yield a small plasmid derivative, pFJ265, that is suitable for Streptomyces gene cloning. pFJ265 is a 9.2-kb nonconjugative plasmid and has a copy number of several hundred per chromosome.