Given current global concerns regarding the accumulation of plastic waste in the environment, biopolymers have become a ‘hot topic’ of research in several fields. However, special attention should be paid to assessing how biodegradable these materials really are – most biopolymers require special conditions to even initiate the degradation process, and very few can be degraded within an acceptable timeframe. Polyhydroxybutyrate (PHB) is a natural biodegradable polyester, enzymatically degraded by a variety of microbial strains which release PHB depolymerases (PhaZs). In this study, we produced five heterologous extracellular PhaZs from different bacterial strains – isolated from various environments like soil and sea water - and compared their degradation capability under different conditions. Recombinant His-tagged modified versions of PhaZs from Comamonas testosteroni, Cupriavidus sp., Marinobacter algicola, Pseudomonas stutzeri, and Ralstonia sp. were successfully expressed in E. coli. PhaZs expression levels along with purity varied for each enzyme; purification protocols were therefore optimized. Activity was assessed by the formation of transparent halos on agar-PHB plates at enzyme concentrations of 2 µg/ml at temperatures ranging from 15°C to 37°C and pH 4.3 to 7. At 37°C, each enzyme displayed similar activity, showing degradation from the first day of incubation. At lower temperatures, each enzyme showed activity after a longer incubation period; nonetheless, at 15°C PhaZ from M. algicola presented higher activity compared to the other PhaZs, which can be explained by its marine origin. At pH 4.3, all enzymes were rendered inactive, but at pH 5.4 PhaZs from C. sp. and R. sp. retained a good level of activity compared to the other PhaZs. These observations present a basis for selecting PhaZs for diverse applications, such as scheduled PHB degradation or biosensors that act under harsh temperature or pH conditions.

Comparative analysis of recombinant polyhydroxybutyrate depolymerases from bacterial strains

254th National Meeting (2017)

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