Page 325 - Demo

SECTION II. APPLIED BIOTECHNOLOGY IN THE PHARMACEUTICAL INDUSTRY... 329The term %u201cribosome engineering%u201d was introduced in 1996 when streptomycin was used to induce a mutation in the rpsLgene, which encodes ribosomal protein S12, in Streptomyces lividans [7]. This mutation, involving the substitution of Lys-88 with Glu, resulted in the activation of actinorhodin production. The nucleotide guanosine 5%u2019-diphosphate 3%u2019-diphosphate (ppGpp) plays a crucial role in responding to nutrient limitations and triggering antibiotic biosynthesis by binding to RNA polymerase. ppGpp is synthesized by ppGpp synthase (RelA), a ribosomal subunit. The streptomycin-induced mutation in ribosomal protein S12 leads to an increase in ppGpp levels, thereby enhancing or activating secondary metabolite production [8,9]. Ribosome engineering has been used to activate antibiotic production in a wide range of actinomycetes using antibiotics like streptomycin, rifampicin, and gentamicin. In tests with different strains, 29% of Streptomyces strains and 8% of non-Streptomyces strains showed activation of secondary metabolite production [10-12].In this study, ribosome engineering was used to enhance the pradimicin A production in A. hibisca P172-2 by using streptomycin.2. MATERIALS AND METHODS 2.1. Strains and culture conditionsA. hibisca P172-2 was cultured in various media for different purposes. For seed culture, ISP2 broth (containing 4 g of yeast extract, 10 g of malt extract, and 4 g of dextrose in 1 L distilled water) was used. For strain maintenance, A. hibisca P172-2 was cultured in ISP2 agar media. For inducing rpsL mutation, MS plate (20 g/L of mannitol, 20 g/L of soya flour, 10 mM of CaCl2, and 20 g/L of agar) were supplemented with 200 %u00b5g/mL of streptomycin. For pradimicin A production, the production media consisted