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Molecular cloning and in silico analysis of chalcone isomerase from Polygonum minus

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Abstract

Background

Chalcone isomerase (CHI; EC 5.5.1.6) is one of the key enzymes in the flavonoid biosynthetic pathway that is responsible for the intramolecular cyclization of chalcones into specific 2S-flavanones.

Methods and results

In this study, the open reading frame (ORF) of CHI was successfully isolated from the cDNA of Polygonum minus at 711-bp long, encoding for 236 amino acid residues, with a predicted molecular weight of 25.4 kDa. Multiple sequence alignment and phylogenetic analysis revealed that the conserved residues (Thr50, Tyr108, Asn115, and Ser192) in the cleft of CHI enzyme group active site are present in PmCHI protein sequence and classified as type I. PmCHI comprises more hydrophobic residues without a signal peptide and transmembrane helices. The three-dimensional (3D) structure of PmCHI predicted through homology modeling was validated by Ramachandran plot and Verify3D, with values within the acceptable range of a good model. PmCHI was cloned into pET-28b(+) plasmid, expressed in Escherichia coli BL21(DE3) at 16 °C and partially purified.

Conclusion

These findings contribute to a deeper understanding of the PmCHI protein and its potential for further characterization of its functional properties in the flavonoid biosynthetic pathway.

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Data availability

Data will be made available upon reasonable request.

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Acknowledgements

Fatin Lyana Azman Shah was supported by a scholarship from Jabatan Perkhidmatan Awam (JPA) Malaysia.

Funding

The work was supported through the Special Allocation for Agencies Under Academy of Science Malaysia-Ministry of Science, Technology and Innovation Grant (PKA0514F004, 6300824) and Universiti Putra Malaysia Putra Graduate Initiative (GP/IPS/2016/9482200).

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Authors and Affiliations

  1. Enzyme and Microbial Technology Research Centre, Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra Malaysia, 43400, Serdang, Malaysia

    Fatin Lyana Azman Shah, Thean Chor Leow, Siti Nurbaya Oslan & Suriana Sabri

  2. Department of Microbiology, Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra Malaysia, 43400, Serdang, Malaysia

    Fatin Lyana Azman Shah & Suriana Sabri

  3. Institute of Systems Biology (INBIOSIS), Universiti Kebangsaan Malaysia, 43600, Bangi, Selangor, Malaysia

    Syarul Nataqain Baharum, Hoe-Han Goh & Ahmad Bazli Ramzi

  4. Department of Cell and Molecular Biology, Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra Malaysia, 43400, Serdang, Malaysia

    Thean Chor Leow

  5. Department of Biochemistry, Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra Malaysia, 43400, Serdang, Malaysia

    Siti Nurbaya Oslan

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All authors listed have made a considerable, direct, and logical contribution to the work and approved it for publication. The experiments were designed by FLAS, SNB, HHG, TCL, ABR, SNO and SS. FLAS and SS drafted the manuscript. All authors discussed the results, edited the manuscript, and approved the final version.

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Correspondence to Suriana Sabri.

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Shah, F.L.A., Baharum, S.N., Goh, HH. et al. Molecular cloning and in silico analysis of chalcone isomerase from Polygonum minus. Mol Biol Rep 50, 5283–5294 (2023). https://doi.org/10.1007/s11033-023-08417-1

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