Bernard Loo1, Karen M. Polizzi2, Javier Chaparro-Riggers3, Janna K. Billy4, and Andreas S. Bommarius4. (1) GIT, IBB 3428, 315 Ferst Dr, Atlanta, GA 30332, (2) School of Chemical and Biomolecular Engineering, Georgia Institute of Technology, 311 Ferst Dr, Atlanta, GA 30332, (3) School of Chemical and Biomolecular Engineering, Georgia Institute of Technology, IBB 3428, 315 Ferst Dr, Atlanta, GA 30332, (4) Georgia Institute of Technology, School of Chemical Engineering, 315 Ferst Drive N.W., Parker H. Petit Biotechnology Institute, Room 3310, Atlanta, GA 30332-0363
The directed evolution of enzymes such as Penicillin G Acylase (PGA) for greater synthetic utility will be reported here. PGA is an industrially important enzyme used in the hydrolysis of penicillin G to 6-aminopenicillanic acid (6-APA) and phenyl acetic acid. In turn, 6-APA is used in the synthesis of other b-lactam antibiotics such as amoxicillin and ampicillin. The engineering of PGA is challenging as it is an 86kDa hetero-dimeric protein and it has a complex maturation process which includes production of a precursor which is eventually exported in the periplasm for processing. The signal sequence from alpha-subunit is removed and the alpha-beta linker region is also removed via intra-molecular proteolysis. We applied semi-rational design on PGA for improved functional properties and will report on our findings. An enhanced PGA would be advantageous for improved green processes for semi-synthetic synthesis of conventional and novel antibiotics. Potentially useful enzymes for antibiotics synthesis will also be discussed.