Preprint Article Version 1 Preserved in Portico This version is not peer-reviewed

Role of the C-terminal β Sandwich of Thermoanaerobacter tengcongensis Thermophilic Esterase in Hydrolysis of Long Chain Acyl Substrates

Enoch B Joel
ORCID logo ,
Adepeju Aberuagba
,
Adebayo Bello
,
Mariam Akanbi-Gada
,
Adedoyin Igunnu
,
Sylvia Omonirume Malomo
,
Femi J. Olorunniji
* ORCID logo
Version 1 : Received: 19 November 2023 / Approved: 20 November 2023 / Online: 20 November 2023 (05:15:28 CET)

A peer-reviewed article of this Preprint also exists.

Joel, E.B.; Aberuagba, A.; Bello, A.J.; Akanbi-Gada, M.; Igunnu, A.; Malomo, S.O.; Olorunniji, F.J. Role of the C-Terminal β Sandwich of Thermoanaerobacter tengcongensis Thermophilic Esterase in Hydrolysis of Long-Chain Acyl Substrates. Int. J. Mol. Sci. 2024, 25, 1272. Joel, E.B.; Aberuagba, A.; Bello, A.J.; Akanbi-Gada, M.; Igunnu, A.; Malomo, S.O.; Olorunniji, F.J. Role of the C-Terminal β Sandwich of Thermoanaerobacter tengcongensis Thermophilic Esterase in Hydrolysis of Long-Chain Acyl Substrates. Int. J. Mol. Sci. 2024, 25, 1272.

Abstract

To search for novel thermostable esterase for optimized industrial applications, esterase from a thermophilic eubacterium species, Thermoanaerobacter tengcongensis MB4 was purified and characterised in this work. Sequence analysis of T. tengcongensis esterase with other homologous esterases of the same family revealed an apparent tail at the C-terminal that is not conserved across the esterase family. Hence, it was hypothesized that the tail is unlikely to have an essential structural or catalytic role. However, there is no documented report of any role for this tail region. We probed the role of the C-terminal domain on the catalytic activity and substrate preference of Thermoanaerobacter tengcongensis esterase EstA3 with a view to see how it could be engineered for enhanced properties. To achieve this, we cloned, expressed, and purified the wild-type and the truncated version of the enzyme. In addition, a naturally occurring member of the family (from B. brevis) that lacks the C-terminal tail was also made. In vitro characterization of the purified enzymes showed that the C-terminal domain contributes significantly to the catalytic activity and distinct substrate preference of T. tengcongensis esterase EstA3. All the three recombinant enzymes showed highest preference to paranitrophenyl butyrate (pNPC4), which suggests they are true esterases, not lipases. kinetic data revealed that truncation had a slight effect on the substrate-binding affinity. Thus, the drop in preference towards long-chain substrates might not be a result of substrate binding affinity alone.

Keywords

Esterase; Thermoanaerobacter tengcongensis; C-terminal domain; substrate preference; catalytic activity

Subject

Biology and Life Sciences, Biology and Biotechnology

Copyright: This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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