A new antibiotic, cypemycin. Taxonomy, fermentation, isolation and biological characteristics.

Abstract

A newpeptide antibiotic, cypemycin, with a molecular weight of 2,097 (M +H), was isolated from the culture broth of Streptomyces sp. OH-4156.The antibiotic possesses cytocidal activity against P388 leukemia cells in vitro at a concentration of 1.3 //g/ml (IC50 values), and the antibiotic showed antimicrobial activities against Micrococcus luteus (MIC, 0.2 /jg/ml).In the course of a screening program for novel antibiotics showing cytocidal activity, cypemycin was isolated from the culture broth of Streptomyces sp. OH-4156 which had been isolated from a soil sample collected in Tokyo, Japan. The antibiotic exhibited cytocidal activity against mammalian tumor cells in vitro, and antimicrobial activity against Micrococcus luteus.The present paper deals with taxonomic studies of the producing strain, and the production, isolation and physico-chemical properties of the new antibiotic. The preliminary biological activities of cypemycin are also described. Materials and Methods General Experimental ProceduresSilica gel 60 (Merck) and Sephadex LH-20 (Pharmacia Fine Chemicals) were used for column chromatography and pre-coated TLCplates of Silica gel 60 (Merck) was used for TLC analysis. Taxonomic StudiesThe type diaminopimelic acid (DAP) was determined by the method of Takahashi et al}\ To investigate the cultural and physiological characteristics, the International Streptomyces Project (ISP) media recommended by Shiriling and Gottlieb2) and those recommended by Waksman3) were used. Cultures were observed after incubation at 27°C for 2 weeks. Color names and hue numbers indicated in Table 1 are those of the Color Harmony Manual (4th Ed.)4). The utilization of carbon sources was tested by growth on Pridham and Gottlieb's medium5) containing a 1%carbon source at 27°C. Cytotoxic Activity TestsSeven strains of established mammaliancells were maintained in monolayers or in suspension in Eagle's minimum essential medium (MEM)supplemented with 10% bovine serum and kanamycin (60 Atg/ml). To determine the cytotoxicity of cypemycin, cells suspended in 200 /zl of the medium were plated in a 96-well culture plate (Falcon) and incubated for 24 hours at 37°C in a 5% CO2-95%air atmosphere. To each well was added 5 fi\ of medium containing a different concentration of cypemycin. After 72 hours of incubation, the cell growth was evaluated by the method of Miravelli et al.6\

Biosynthesis and Regulation of Grisemycin, a New Member of the Linaridin Family of Ribosomally Synthesized Peptides Produced by Streptomyces griseus IFO 13350

“…It was speculated that the greater susceptibility to pore formation of micrococci than that of lactobacilli could reflect, in part, their relatively thinner phospholipid bilayer. The lack of antimicrobial activity observed for grisemycin and the very narrow spectrum reported for cypemycin (11) suggest that these two compounds might serve different roles in the biology of their producing Streptomyces organisms, for example, as signaling molecules.…”

“…The resulting S. griseus ⌬grmA::[oriT-aac(3)IV] mutant strain was designated M1458. For bioassays, strains were grown for 3 days in cypemycin production medium (11), and culture supernatants were separated from the mycelium by centrifugation and extracted with CHCl 3 . The solvent was evaporated and the residual pellet dissolved in 5% formic acid for matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) analysis or in methanol (MeOH) for use in an M. luteus paper disc bioassay (3).…”

Genome mining and genetic analysis of cypemycin biosynthesis reveal an unusual class of posttranslationally modified peptides

“…In this study, we identified the cypemycin biosynthetic gene cluster of Streptomyces sp. OH-4156 (7) and show that these previously described modifications are introduced by unusual enzymes or by an unusual modification pathway. Six distinct posttranslational modifications participate in cypemycin biosynthesis, altering 9 of the 22 amino acids found in the mature peptide.…”

Lantibiotics produced by Actinobacteria and their potential applications (a review)

“…Actinobacterial bacteriocins inhibit the growth of species related to the producer, such as michiganin A [13], although many of them possess a broad spectrum of inhibitory activity that includes non-related bacteria (microbisporicin), protozoans (thiostrepton), yeasts and fungi (cinnamycin) and viruses (labyrinthopeptin A1 and duramycin) [14][15][16][17][18]. In addition to antimicrobial activity, some actinobacterial bacteriocins and their semisynthetic derivatives also exhibit anti-inflammatory [19][20][21], anti-allergic [20], antitumour [22][23][24] and antinociceptive activities [25,26], and may also act in blood pressure regulation [19,27]. Some of these peptides have already found clinical applications, such as thiostrepton, a thiopeptide used in dermatologic ointment for veterinary use [28], and others are in stages of pre-clinical and clinical trials, such as NAI-107, NAI-112 [29] and lancovutide/duramycin (https://clinicaltrials.gov/ct2/show/study/ NCT00671736?term=Lancovutide&rank=1; accessed September 17,2016), which reinforces the great potential of practical application of bacteriocins produced by Actinobacteria.…”