Preparation of cross-linked enzyme aggregates in water-in-oil emulsion: Application to trehalose synthase

• Spherical CLEA of trehalose synthase was prepared in a water-in-oil emulsion.
• Particle size of spherical CLEA is 20–60 μm in diameter.
• Spherical CLEA was more tolerant to metal ions and denaturants than the free enzyme.

In this work, a novel method to prepare cross-linked enzyme aggregates (CLEA) in a water-in-oil emulsion was reported. Spherical CLEA of recombinant trehalose synthase (TreS) from Meiothermus rubber was obtained through the emulsion based process. By varying the experimental parameters, the optimum conditions for the preparation of spherical CLEA were determined. The optimum catalytic temperature and thermal stability of spherical CLEA were enhanced significantly compared with the free enzyme. Spherical CLEA was also more tolerant to metal ions and chemical denaturants. Optical and scanning electron microscopy analysis showed that instead of the amorphous clumping shape of conventionally prepared CLEA, spherical CLEA has a more structured ball-like appearance with a size of 20–60 μm in diameter that could be easily separated.

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