Studies on lipase from aeromonas veronii PG01

Abstract

Lipases (Carboxyl ester hydrolases EC.3.1.1.3) are enzymes which hydrolyse long chain fatty acid esters. Lipases also catalyse the reverse reaction from glycerol and fatty acids to form glycerides by esterification, thus lipases are active in both aqueous and nonaqueous solvent systems. The first objective of the present work is the isolation and characterization of organic solvent tolerant bacteria that produces organic solvent-stable extracellular lipase. The second objective of this work is purification and characterization of the organic solvent stable lipase. Third objective is identification of substrate binding domain of lipase in Aeromonas veronii PG01. The second objective of this work the A.veronii PG01lipase was purified to approximately 4 fold with a specific activity of 1240 U/mg and an overall yield of 20% by a combination of ammonium sulfate precipitation, Hydrophobic interaction chromatography and ion exchange chromatography. We could not get maximum recovery of lipase after two step purification as most of the lipase was lost during purification step. The third objective of this work was to find out the putative lipase substrate binding domain of Aeromonas veronii PG01. PCR amplified product was subcloned into the pGEM-T vector. After lysate PCR reaction the positive colony was used for DNA sequencing reaction. Generally, all Aeromonas sp. contains (V-H-F-L-G-H-S-L-G-A) amino acid residue in substrate binding domain whereas, the nucleotide sequence of corresponding aminoacids are varied within the genus. Aeromonas veronii PG01 lipase substrate binding domain nucleotide sequences are similar to A.sobria lipase substrate binding domain sequence. newline newline newline