PEMURNIAN DAN KARAKTERISASI ENZIM-PGA INTRASEL DARI BACILLUS SP. BAC4

• Main Author: Bahri, Syaiful
• Format: Theses
• Language: Indonesia
• Online Access: https://digilib.itb.ac.id/gdl/view/2263
• Institution: Institut Teknologi Bandung

A local strain of Bacillus sp. BAC4 has been identified to produce penicillin G acylase (PGA). This enzyme hydrolyses benzylpenicillin to 6-aminopencillanic acid and phenylacetic acid. The 6-aminopenicillanic acid is a valuable starting material for the production of various derivatives of (3-laktam antibiotics. Previous research had shown that Bacillus sp. BAC4 able to produce both extracellular and intracellular PGA. The extracellular PGA of Bacillus sp. BAC4 has been studied extensively, but the intracellular PGA has not been characterized. This research was aimed to study the properties of intracellular PGA, through purification and characterization. Intracellular PGA purification was done by sonication, followed by centifugation to sparated the filtrate from cell debris. The filtrate was than fractionated by 80 % (w/v) ammonium sulphate, dialysed in selophan tube, and fractionated by affinity chromatography using phenylacetate ligands. Three fractions were obtained, two of which showed protease activity whilst the other one has acylase activity. Acylase fraction contained a protein of 32.5 mg with a specific activity of 0.29 unit/mg. Further characterization by SDSPAGE showed two distinct bands at 47 kD and 35 kD. This data is in in good agreement with the intracellular PGA of E. coil, K citrophilia, A. viscocus and P. rettgeri which also contained of two sub-unit moleculs with a relative molecular mass of 80 kD. This low molecular weight was unexpected, as the extracellular PGA of Bacillus sp. BAC4 having a larger molecular mass of 130 kD. This result suggest that the intracellular and the extracellular PGA of Bacillus sp. BAC4 might be of two different uncorrelated molecules.