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Esterification of Polymeric Carbohydrate Through Congener Cutinase-Like Biocatalyst

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Abstract

Cutinase-like enzymes (CLEs) are bi-functional hydrolases, which share the conserved catalytic site of lipase and consensus pentapeptide sequence of cutinase. Here, we have genetically replaced the canonical amino acids (CAA) by their non-canonical fluorinated surrogates to biosynthesize a novel class of congener biocatalyst for esterification of polymeric carbohydrate with long-chain fatty acid. It is a new enzyme-engineering approach used to manipulate industrially relevant biocatalyst through genetic incorporation of new functionally encoded non-canonical amino acids (NCAA). Global fluorination of CLE improved its catalytic, functional, and structural stability. Molecular docking studies confirmed that the fluorinated CLE (FCLE) had developed a binding affinity towards different fatty acids compared with the parent CLE. Importantly, FCLE could catalyze starch oleate synthesis in 24 h with a degree of substitution of 0.3 ± 0.001. Biophysical and microscopic analysis substantiated the efficient synthesis of the ester by FCLE. Our data represent the first step in the generation of an industrially relevant fluorous multifunctional enzyme for facile synthesis of high fatty acid starch esters.

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Acknowledgments

We thank the Director, CSIR-CLRI, for his support during the project. We also thank ICMR (File No: 5/3/8/4/2019-ITR) for research support.

Funding

We acknowledge the Council of Scientific and Industrial Research (CSIR), New Delhi, for the award of Senior Research Fellowship to Ms. Sisila V and the Indian Council of Medical Research (ICMR) for Research Associate to Dr. M. Aarthy. The authors are grateful to CSIR-CLRI for funding this research through the “Major Laboratory Project - Biotechnology division.”

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Authors and Affiliations

  1. Division of Biochemistry and Biotechnology, Council of Scientific and Industrial Research (CSIR), Central Leather Research Institute (CLRI), Chennai, India

    Valappil Sisila, Puhazhendi Puhazhselvan, Mayilvahanan Aarthy, Numbi Ramudu Kamini & Niraikulam Ayyadurai

  2. Academy of Scientific and Innovative Research, Ghaziabad, India

    Valappil Sisila, Numbi Ramudu Kamini & Niraikulam Ayyadurai

  3. Department of Biotechnology, Rajalakshmi Engineering College, Chennai, India

    Girimanikandan Sakkeeshyaa & Perisamy Saravanan

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  1. Valappil Sisila
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  2. Puhazhendi Puhazhselvan
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Correspondence to Niraikulam Ayyadurai.

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Sisila, V., Puhazhselvan, P., Aarthy, M. et al. Esterification of Polymeric Carbohydrate Through Congener Cutinase-Like Biocatalyst. Appl Biochem Biotechnol 193, 19–32 (2021). https://doi.org/10.1007/s12010-020-03415-6

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