P175: Process and strain development for production of novel antibacterial compound PM181104

Screening of natural product libraries against multi drug resistant bacteria such as MRSA and VRE resulted in the discovery of novel thiopeptide antibiotic PM181104 from marine actinobacterium (Patent WO/2007/119201). PM181104 showed potent antibacterial activity against MRSA and VRE through in-vivo studies, hence selected as a lead candidate for drug development. The production of PM181104 to cater the needs of the pre-clinical and toxicological studies was a great challenge since the yield at the discovery stage was very poor. The producer strain, Kocuria spp. produced less than 5mg/L of this antibiotic on Zobell Marine Broth medium in shake flask. Hence a new soya peptone based vegetative medium was formulated that produced 18mg/L of activity in shake flask. Large-scale production of PM181104 was carried out in 30KL capacity fermentor with average titer of 39.87 mg/L using optimized conditions. In order to improve the strains potential to produce PM181104 antibiotic, the strain improvement platform technology was established consisting of classical mutagenesis, rational selection and screening of mutants. Antibiotic resistance was used as rational selection criteria for mutant selection followed by screening in shake flask. Through screening, mutant UV30Neo45(24) having resistance to neomycin (45µg/ml) was found to produce 37 mg/L of antibiotic in shake flask which is about  seven fold improvement than the parent strain.  The mutant strain was further characterized by RAPD and SEM to differentiate it from the parent strain. Further rounds of strain improvement combined with metabolic engineering are underway as a step towards commercialization of this novel antibiotic in future.