PENGHASILAN, PENULENAN DAN PENCIRIAN ENZIM DEPOLIMERASE POLIHIDROKSIALKANOAT BERANTAISEDERHANA DARIPADA PSEUDOMONAS sp.
Loading...
Date
2005-07
Authors
WAN CHIK, MOHD HAYAZI
Journal Title
Journal ISSN
Volume Title
Publisher
Abstract
Twenty six bacterial isolates with the ability to degrade medium chain length
polyhydroxyalkanoates (MCL PHA), were obtained from several areas in Pulau Pinang,
Malaysia. In this study, three methods have been carried out to isolate local MCL PHA
depolymerase producer, namely the screening method using isolation medium containing
MCL PHA as the sole carbon source, the degradation index method and the enzyme
assay method. Among these isolates, NT-Dp B which showed the best ability to produce
MCL PHA was selected for further investigation. Identification test showed that the
bacterium was Pseudomonas sp. The bacterium was unable to degrade short chain length
polyhydroxyalkanoates (SCL PHA) such as poly(3-hydroxybutyrate) [P(3HB)] and
poly(3-hydroxybutyrate-co-3-hydroxyvalerate), [P(3HB-co-3HV)]. Further studies on the
bacteria ability to produce PHA showed that the bacteria was capable of accumulating
MCL PHA from carbon sources such as glucose, 4-hydroxybutyric acid, succinate and
oleic acid. Pseudomonas sp. was able to produce extracellular depolymerase capable of
degrading MCL PHA. After optimizing the media composition and culture condition
using shake flask, the enzyme activity obtained was 0.749 mg/mUminutes, which was 20
fold higher compared to before the optimization process. The medium composition for
optimized production of the depolymerase enzyme consisted of 2.5 giL MCL PHA, 6.4
XXIV
giL urea, 4.0 giL KH2P04, 2,5 giL K2HP04, 1.0 giL MgS04.1H20, 3% (v/v) ferric
ammonium citrate and calcium chloride solution. Whereas, the physical parameters for
optimized depolymerase production were culture temperature at 30°C, pH medium at 6.8,
inoculums size of 0.030 giL and agitation at 200 rpm. The purification process was
successfully carried out and separated the enzymes from other kind of proteins. The
purified depolymerase enzyme had molecular weight of 22.4 kDa and isoelectric point
was at pH 8.8. The enzyme showed optimum activity at 40°C and was stable from 30°C
to 35°C. The optimum pH was 8.5, and the enzyme was stable at pH 9.5 in glycineNaOH
buffer. The enzyme Km was 0.74 mg/ml whereas the Vmaxs was 0.27
mg/mL/minutes.
Description
Keywords
PENGHASILAN, PENULENAN DAN PENCIRIAN ENZIM DEPOLIMERASE POLIHIDROKSIALKANOAT BERANTAISEDERHANA