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In silico characterization and differential expression analysis of 1-deoxy-d-xylulose-5-phosphate reductoisomerase (DXR) of Centella asiatica

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Abstract

The 1-deoxy-d-xylulose-5-phosphate reductoisomerase (DXR; EC1.1.1.267), an NADPH-dependent reductase, plays a pivotal role in the methylerythritol 4-phosphate pathway (MEP), in the conversion of 1-deoxy-d-xylulose-5-phosphate (DXP) into MEP. Photochemical profiles, as well as pharmaceutical activities of Centella asiatica (L.), one of the most valuable medicinal plants, divulge the presence of secondary metabolites called Centellosides. Despite well-studied pharmaceutical activities, not much is known about the genes responsible for the synthesis of these compounds. In the present study, the full-length DXR gene sequence (JQ965955) of Centella submitted in NCBI was characterized using various bioinformatics tools and tissue specific differential expression studies were also carried out. The full-length CDNA of CaDXR contains an open reading frame (ORF) of 1425 bp which encodes a peptide of 474 amino acids. The molecular weight of this protein was found to be 51.5 kDa with isoelectric point of 6.33. The protein contains three conserved domain, namely NADPH (GSTGSIGT and LAAGSNV), substrate binding (LPADSEHSAI and NKGLEVIEAHY) and Cys-Ser-(Ala/Met/Val/Thr) cleavage-site domains. Phylogenetic studies of CaDXR sequence show close homology with DXR sequence of Angelica sinensis and Daucus carota subsp sativus as they all belong to Apiaceae family. In silico analysis predicted that CaDXR protein contains 21 α-helix and 11 β-sheets and further DXR protein model was validated by Ramachandran plot analysis. The results of molecular dynamics (MD) simulations unveil dynamic stability of the proposed model and docking studies suggest that the NDP cofactor tightly binds in the active site of the protein with a strong network of hydrogen and hydrophobic interactions. The expression studies by semi-RT followed by qRT—PCR suggests that CaDXR is differentially expressed in different tissues (with maximal expression in the node and lowest in the roots). Thus, characterization and structure–function analysis of DXR gene in Centella facilitate us to understand not only the functions of DXR gene but also regulatory mechanisms involved in the MEP pathway in C. asiatica plant at the molecular level.

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Authors and Affiliations

  1. Department of Agricultural Biotechnology, Assam Agricultural University, Jorhat, 785013, India

    Richa Sharma, Kamalakshi Devi, Mahendra K. Modi & Priyabrata Sen

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  1. Richa Sharma
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  2. Kamalakshi Devi
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  3. Mahendra K. Modi
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  4. Priyabrata Sen
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Contributions

RS conceived the presented idea and performed the computation. KD investigated and verified the computed data. MKM helped supervised the project. PS contributed to the interpretation of the results and took the lead in writing the manuscript. All authors discussed the results and contributed to the final manuscript.

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Correspondence to Priyabrata Sen.

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We wish to confirm that there are no known conflicts of interest associated with this publication and there has been no significant financial support for this work that could have influenced its outcome.

Supplementary Information

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13205_2021_2723_MOESM1_ESM.tif

Fig.S1 Multiple sequence alignment of CaDXR sequence with all the selected plant DXR sequences using MEGA v6.1 and visualized in ESPript. Conserved regions are highlighted in blue square boxes and labeled in black namely M-I &M-II: NADPH-binding motif, M-III &IV: Substrate-binding motif and M-V: Cleavage-site motif (TIF 632 KB)

13205_2021_2723_MOESM2_ESM.tif

Fig. S2 The molecular model of CaDXR protein was obtained using galaxy web server. The model shows N-terminal β-sheets and C-terminal end of the protein contain α-helix. Structure of the DXR protein of Centella predicted 21 α-helix and 11 β-sheets. (TIF 38747 KB)

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Sharma, R., Devi, K., Modi, M.K. et al. In silico characterization and differential expression analysis of 1-deoxy-d-xylulose-5-phosphate reductoisomerase (DXR) of Centella asiatica. 3 Biotech 11, 184 (2021). https://doi.org/10.1007/s13205-021-02723-w

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