Brazilian Journal of Microbiology
Print version ISSN 1517-8382
CARNEIRO-DA-CUNHA, Maria das Graças; LIMA FILHO, José Luiz de and CAMPOS-TAKAKI, Galba Maria de. Protoplast formation and regeneration from Streptomyces clavuligerus NRRL 3585 and clavulanic acid production. Braz. J. Microbiol. [online]. 2002, vol.33, n.4, pp. 347-351. ISSN 1517-8382. http://dx.doi.org/10.1590/S1517-83822002000400014.
Protoplasts of the wild type Streptomyces clavuligerus NRRL 3585 (ATCC 27064) were formed from spores cultures obtained in the lag, exponential and stationary growth phases by using 0.5% glycine in the culture medium. The protoplasts were obtained by treatment of the cells with lysozyme (EC-220.127.116.11) 40,000 U (1mg/mL), in an osmotic solution for 90 min at 28ºC. The frequency of regenerated protoplasts in the lag phase was 1.7x103 CFU/mL (28.97%), in the beginning of the exponential phase 0.4x102 CFU/mL (31.67%), in the exponential growth phase 2.5x103 CFU/mL (46.30%) and 1.0x105 CFU/mL in stationary phase (48.45%). Antibiotic production and activity of regenerated protoplasts were observed in all phases, except in the lag phase. The protoplast formation and regeneration techniques resulted in a new isolate strain of Streptomyces clavuligerus that produced approximately 2.5 fold more clavulanic acid.
Keywords : Streptomyces clavuligerus; protoplast; clavulanic acid.