TITLE:
An Approach to the Identification and Characterisation of a Psychrotrophic Lipase Producing Pseudomonas sp ADT3 from Arctic Region
AUTHORS:
Arpita Dey, Amarnath Chattopadhyay, Pradipta Saha, Subhrakanti Mukhopadhyay, Tushar Kanti Maiti, Sabyasachi Chatterjee, Pranab Roy
KEYWORDS:
Psychrotrophic; Arctic; Extracellular Lipase; 16S rRNA; Mass Spectrometry
JOURNAL NAME:
Advances in Bioscience and Biotechnology,
Vol.5 No.4,
March
12,
2014
ABSTRACT: The aim of this
research work was to explore psychrotrophic microbes from soil sample of NyAlesund, Svalbard, arctic region and to investigate their potential use as an
effective tool for industrial application. A novel psychrotrophic bacterial
strain showed good growth on minimal medium containing lipid as the only carbon
source. Microbiological characterisation of the isolate showed that it was a
gram negative rod. The strain was tested for the production of extracellular
lipase enzyme. The enzymes were partially purified by 90% saturated ammonium
sulfate and dialysis for desalting. The bacterium was
identified as Pseudomonas sp ADT3 by 16S rRNA amplification and
sequencing which had been deposited in the NCBI GenBank with accession number
JX914667. Phylogenetic tree was also constructed with MEGA5 software and showed
the highest level of sequence similarity with Pseudomonas sp. HC3-13 strain. The microorganism had a growth
optimum at pH 8.0 and temperature 22°C.
Optimization of different parameters e.g. temperature, pH, incubation time,
cofactors etc. was performed for the extracellular lipase activity. The hydrolytic
activity of the enzyme was enhanced 5 times by Pb2+ but strongly
inhibited by heavy metals Hg2+ as well as EDTA and β-mercaptoethanol. For the molecular
weight estimation of enzyme SDS-PAGE was done which showed an inducible band of
approximately 13.9 KDa. Activity staining and mass spectrometry techniques
were also performed.