Enzymatic degradation of polyhydroxyalkanoate using lipase from Bacillus subtilis

Polyhydroxyalkanoates (PHAs) are an important class of biodegradable polymers synthesized by a few bacteria under nutrient-limiting conditions. In this study, the lipase-catalysed degradation of PHA synthesized by Enterobacter sp. was monitored. For this, the lipase-encoding gene from Bacillus subtilis DI2 was PCR-amplified, cloned into a T vector system and sequenced. It was expressed in Escherichia coli DH5 alpha cells, the recombinant enzyme was purified 24.25-fold, and its molecular weight was determined to be around 28 kDa. When PHA biodegradation studies were carried out with this enzyme, gel permeation chromatography showed 21.3 and 28.3 % molecular weight decrease and weight loss, respectively. Further, scanning electron micrographs revealed alterations in polymer surface morphology. Changes in molecular vibrations were noticed in the FTIR spectra. When the chemical shifts in NMR spectra were studied, a steep reduction in area under the peak at 1.57 ppm was observed. In the heating range of 30-930 degrees C employed during thermogravimetry analysis, the degraded sample showed a total of 45.82 % weight loss, as against 18.89 % for the native sample. The melting temperature (T-m) of the polymer was also brought down from 126.22 to 118.18 degrees C, as inferred from differential scanning calorimetry. Lipase-catalysed chain scission reactions could thus be used to generate low molecular weight functional biopolymers with wide-ranging pharmaceutical applications, such as in sustained drug release.