Cloning, protein expression and biochemical characterization of Carica papaya esterase

Abstract

Background

GDSL-like esterase/lipase proteins (GELPs) are enzymes that possess unique characteristics, they contain four invariable catalytic residues. Advances in the study of these proteins are interesting. The cloning and functional expression of a papaya esterase have not been reported. Therefore, in this work we evaluated the heterologous production of Carica papaya esterase CpEST in the yeast Komogataella phaffii (Pichia pastoris).

Results

The cloning and expression of the protein was performed under the P_AOX1 promoter, and productions of up to 43 AU/mL were achieved using residual glycerol from biodiesel in the batch phase and methanol for the induction phase. Enzyme activity assays determined that CpEST has a high preference for short-chain substrates (p-NP C4 and p-NP C8), and optimal activity conditions were observed at 30°C and pH 10. The enzyme showed the highest stability to acetone, ethanol and tert-butanol solvents, retaining approximately 55% of its initial enzymatic activity after 1 h of exposure.

Conclusions

Cloning and functional expression of papaya CpEST esterase was achieved. During fermentation, the yeasts used as a carbon source residual glycerol from biodiesel production. Based on the results obtained from the characterization of the esterase, it was found that it has a high potential for use in the bioenergy and detergent industry.

How to cite: Reyes-Reyes AL, Valero F, Sandoval G. Cloning, protein expression and biochemical characterization of Carica papaya esterase. Electron J Biotechnol 2023;61. https://doi.org/10.1016/j.ejbt.2022.11.004.