Skip to main content
SpringerLink
Log in

Enhancement of L-ribulose Production from L-ribose Through Modification of Ochrobactrum sp. CSL1 Ribose-5-phosphate Isomerase A

  • Original Article
  • Published:
Applied Biochemistry and Biotechnology Aims and scope Submit manuscript

Abstract

L-ribulose, a kind of high-value rare sugar, could be utilized to manufacture L-form sugars and antiviral drugs, generally produced from L-arabinose as a substrate. However, the production of L-ribulose from L-arabinose is limited by the equilibrium ratio of the catalytic reaction, hence, it is necessary to explore a new biological enzymatic method to produce L-ribulose. Ribose-5-phosphate isomerase (Rpi) is an enzyme that can catalyze the reversible isomerization between L-ribose and L-ribulose, which is of great significance for the preparation of L-ribulose. In order to obtain highly active ribose-5-phosphate isomerase to manufacture L-ribulose, ribose-5-phosphate isomerase A (OsRpiA) from Ochrobactrum sp. CSL1 was engineered based on structural and sequence analyses. Through a rational design strategy, a triple-mutant strain A10T/T32S/G101N with 160% activity was acquired. The enzymatic properties of the mutant were systematically investigated, and the optimum conditions were characterized to achieve the maximum yield of L-ribulose. Kinetic analysis clarified that the A10T/T32S/G101N mutant had a stronger affinity for the substrate and increased catalytic efficiency. Furthermore, molecular dynamics simulations indicated that the binding of the substrate to A10T/T32S/G101N was more stable than that of wild type. The shorter distance between the catalytic residues of A10T/T32S/G101N and L-ribose illuminated the increased activity. Overall, the present study provided a solid basis for demonstrating the complex functions of crucial residues in RpiAs as well as in rare sugar preparation.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7
Fig. 8
Fig. 9

Similar content being viewed by others

References

  1. Zhang, W. L., Zhang, T., Jiang, B., & Mu, W. M. (2017). Enzymatic approaches to rare sugar production. Biotechnology Advances, 35, 267–274.

    Article  Google Scholar 

  2. Chen, D., Chen, J., Liu, X., Guang, C., Zhang, W., & Mu, W. (2021). Biochemical identification of a hyperthermostable L-ribulose 3-epimerase from Labedella endophytica and its application for D-allulose bioconversion. International Journal of Biological Macromolecules, 189, 214–222.

    Article  CAS  Google Scholar 

  3. Mahmood, S., Iqbal, M. W., Riaz, T., Zhang, W. L., & Mu, W. M. (2020). Characterization of recombinant L-ribose isomerase acquired from Cryobacterium sp. N21 with potential application in L-ribulose production. Process Biochemistry, 97, 1–10.

    Article  CAS  Google Scholar 

  4. Hu, C., Li, L. Z., Zheng, Y. Y., Rui, L. L., & Hu, C. Y. (2011). Perspectives of biotechnological production of l-ribose and its purification. Appl Microbiol Biot, 92, 449–455.

    Article  CAS  Google Scholar 

  5. Okano, K. (2009). Synthesis and pharmaceutical application of L-ribose. Tetrahedron, 65, 1937–1949.

    Article  CAS  Google Scholar 

  6. Guo, Z. R., Long, L. K., & Ding, S. J. (2019). Characterization of a D-lyxose isomerase from Bacillus velezensis and its application for the production of D-mannose and L-ribose. Amb Express, 9, 149–160.

    Article  Google Scholar 

  7. Helanto, M., Kiviharju, K., Leisola, M., & Nyyssola, A. (2007). Metabolic engineering of lactobacillus plantarum for production of L-ribulose. Applied and Environmental Microbiology, 73, 7083–7091.

    Article  CAS  Google Scholar 

  8. Jiajun, C., Hao, W., Wenli, Z., & Wanmeng, M. (2020). Recent advances in properties, production, and applications of L-ribulose. Applied Microbiology and Biotechnology, 104, 5663–5672.

    Article  Google Scholar 

  9. Mahmood, S., Iqbal, M. W., Zhang, W. L., & Mu, W. M. (2021). A review on L-ribose isomerases for the biocatalytic production of L-ribose and L-ribulose. Food Research International, 145, 110409–110421.

    Article  CAS  Google Scholar 

  10. Ahmed, Z., Shimonishi, T., Bhuiyan, S. H., Utamura, M., Takada, G., & Izumori, K. (1999). Biochemical preparation of L-ribose and L-arabinose from ribitol: A new approach. Journal of Bioscience and Bioengineering, 88, 444–448.

    Article  CAS  Google Scholar 

  11. Helanto, M., Kiviharju, K., Granstrom, T., Leisola, M., & Nyyssola, A. (2009). Biotechnological production of L-ribose from L-arabinose. Appl Microbiol Biot, 83, 77–83.

    Article  CAS  Google Scholar 

  12. Granstrom, T. B., Takata, G., Tokuda, M., & Izumori, K. (2004). Izumoring: A novel and complete strategy for bioproduction of rare sugars. Journal of Bioscience and Bioengineering, 97, 89–94.

    Article  Google Scholar 

  13. Mahmood, S., Iqbal, M. W., Riaz, T., Hassanin, H. A. M., Zhu, Y. Y., Ni, D. W., & Mu, W. M. (2020). Characterization of a recombinant L-ribose isomerase from Mycetocola miduiensis and its application for the production of L-ribulose. Enzyme and Microbial Technology, 135, 109510–109519.

    Article  CAS  Google Scholar 

  14. Liu, X. X., Li, Z. J., Chen, Z., Wang, N., Gao, Y. H., Nakanishi, H., & Gao, X. D. (2019). Production of L-ribulose using an encapsulated L-arabinose isomerase in yeast spores. Journal of Agricultural and Food Chemistry, 67, 4868–4875.

    Article  CAS  Google Scholar 

  15. Yeom, S. J., Kim, N. H., Yoon, R. Y., Kwon, H. J., Park, C. S., & Oh, D. K. (2009). Characterization of a mannose-6-phosphate isomerase from Geobacillus thermodenitrificans that converts monosaccharides. Biotechnology Letters, 31, 1273–1278.

    Article  CAS  Google Scholar 

  16. Ju, X., Xu, X. Q., Shen, M., Mo, X. B., Fan, H., & Li, L. Z. (2020). Biochemical and structural insights into an Ochrobactrum sp. CSL1 ribose-5-phosphate isomerase A and its roles in isomerization of rare sugars. Enzyme and Microbial Technology, 140, 109604–109613.

    Article  CAS  Google Scholar 

  17. Chen, J. J., Wu, H., Zhang, W. L., & Mu, W. M. (2020). Ribose-5-phosphate isomerases: Characteristics, structural features, and applications. Applied Microbiology and Biotechnology, 104, 6429–6441.

    Article  CAS  Google Scholar 

  18. Essenberg, M. K., & Cooper, R. A. (1975). Two ribose-5-phosphate isomerases from Escherichia coli K12: Partial characterisation of the enzymes and consideration of their possible physiological roles. European journal of biochemistry, 55, 323–332.

    Article  CAS  Google Scholar 

  19. Shen, M., Ju, X., Xu, X. Q., Yao, X. M., Li, L. Z., Chen, J. J., Hu, C. Y., Fu, J. L., & Yan, L. S. (2018). Characterization of Rihnse-5-Phosphate Isomerase B from newly isolated strain Ochrobactrum sp CSL1 producing L-rhamnulose from L-rhamnose. Journal of Microbiology and Biotechnology, 28, 1122–1132.

    Article  CAS  Google Scholar 

  20. Wang, R., Xu, X. Q., Yao, X. M., Tang, H. T., Ju, X., & Li, L. Z. (2021). Enhanced isomerization of rare sugars by ribose-5-phosphate isomerase A from Ochrobactrum sp. CSL1. Enzyme and Microbial Technology, 148, 109789–109797.

    Article  CAS  Google Scholar 

  21. Sedmak, J. J., & Grossberg, S. E. J. A. B. (1977). A rapid, sensitive, and versatile assay for protein using coomassie brilliant blue G250. Analytical Biochemistry, 79, 544–552.

    Article  CAS  Google Scholar 

  22. Bradford, M. M. (1976). A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Analytical biochemistry, 72, 248–254.

    Article  CAS  Google Scholar 

  23. Muniruzzaman, S., Tokunaga, H., & Izumori, K. (1994). Isolation of enterobacter agglomerans strain 221E from soil, a potent D-tagatose producer from galactitol. Journal of Fermentation and Bioengineering, 78, 145–148.

    Article  CAS  Google Scholar 

  24. Jung, C. H., Hartman, F. C., Lu, T. Y. S., & Larimer, F. W. (2000). D-ribose-5-phosphate isomerase from spinach: Heterologous overexpression, purification, characterization, and site-directed mutagenesis of the recombinant enzyme. Archives of Biochemistry and Biophysics, 373, 409–417.

    Article  CAS  Google Scholar 

  25. Ishikawa, K., Matsui, I., Payan, F., Cambillau, C., Ishida, H., Kawarabayasi, Y., Kikuchi, H., & Roussel, A. (2002). A hyperthermostable D-ribose-5-phosphate isomerase from Pyrococcus horikoshii characterization and three-dimensional structure. Structure, 10, 877–886.

    Article  CAS  Google Scholar 

  26. Capriles, P., Baprista, L. P. R., Guedes, I. A., Guimaraes, A. C. R., Custodio, F. L., Alves-Ferreira, M., & Dardenne, L. E. (2015). Structural modeling and docking studies of ribose 5-phosphate isomerase from Leishmania major and Homo sapiens: A comparative analysis for Leishmaniasis treatment. Journal of Molecular Graphics & Modelling, 55, 134–147.

    Article  CAS  Google Scholar 

  27. Zhang, X. F., Xu, X. Q., Yao, X. M., Wang, R., Tang, H. T., Ju, X., & Li, L. Z. (2020). Exploring multifunctional residues of ribose-5-phosphate isomerase B from Ochrobactrum sp. CSL1 enhancing isomerization of D-Allose. Journal of Agricultural and Food Chemistry, 68, 3539–3547.

    Article  CAS  Google Scholar 

  28. Kaur, P. K., Tripathi, N., Desale, J., Neelagiri, S., Yadav, S., Bharatam, P. V., & Singh, S. (2016). Mutational and structural analysis of conserved residues in ribose-5-phosphate isomerase B from Leishmania donovani: Role in substrate recognition and conformational stability. Plos One, 11, 0150764–0150784.

    Google Scholar 

  29. Ziwei, C., Wei, X., Wenli, Z., Tao, Z., Bo, J., & Wanmeng, M. (2018). Characterization of a thermostable recombinant L-rhamnose isomerase from Caldicellulosiruptor obsidiansis OB47 and its application for the production of L-fructose and L-rhamnulose. Journal of the Science of Food and Agriculture, 98, 2184–2193.

    Article  Google Scholar 

  30. Wen, L. Q., Zang, L. L., Huang, K., Li, S. S., Wang, R. L., & Wang, P. G. (2016). Efficient enzymatic synthesis of L-rhamnulose and L-fuculose. Bioorganic & Medicinal Chemistry Letters, 26, 969–972.

    Article  CAS  Google Scholar 

Download references

Funding

This study was funded by the National Natural Science Foundation of China (Grant Nos. 21676173 and 32001634).

Author information

Author notes

  1. Qian Zheng and Rong Wang contributed equally to this work.

Authors and Affiliations

  1. School of Chemistry and Life Sciences, Suzhou University of Science and Technology, No. 99 Xuefu Rd., Huqiu District, Suzhou, Jiangsu Province, 215009, People’s Republic of China

    Qian Zheng, Rong Wang, Xin Ju, Xujing Gu, Zhi Chen & Liangzhi Li

  2. Fujian Key Laboratory of Marine Enzyme Engineering, College of Biosciences and Engineering, Fuzhou University, Fuzhou, 350116, People’s Republic of China

    Xinqi Xu

Authors
  1. Qian Zheng
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Rong Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Xin Ju
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Xujing Gu
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Xinqi Xu
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Zhi Chen
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Liangzhi Li
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

Qian Zheng: Writing (original draft) and investigation. Rong Wang: Methodology and investigation. Xin Ju: Conceptualization, methodology, and writing—original draft. Xujing Gu: Investigation. Xinqi Xu: Investigation and writing—original draft. Zhi Chen: Investigation. Liangzhi Li: Project administration, supervision, and writing—review and editing.

Corresponding authors

Correspondence to Zhi Chen or Liangzhi Li.

Ethics declarations

Ethics Approval

No related ethical issues.

Consent to Participate

The authors promise that the work described has not been published previously, that it is not under consideration for publication elsewhere, and that its publication is approved by all authors and tacitly or explicitly by the responsible authorities where the work was carried out.

Consent for Publication

The authors promise that if the manuscript is accepted, it will not be published elsewhere in the same form, in English, or in any other language, without the written consent of the publisher.

Competing Interests

The authors declare no competing interests.

Additional information

Publisher’s Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Supplementary Information

Below is the link to the electronic supplementary material.

Supplementary file1 (DOCX 445 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Zheng, Q., Wang, R., Ju, X. et al. Enhancement of L-ribulose Production from L-ribose Through Modification of Ochrobactrum sp. CSL1 Ribose-5-phosphate Isomerase A. Appl Biochem Biotechnol 194, 4852–4866 (2022). https://doi.org/10.1007/s12010-022-04015-2

Download citation

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s12010-022-04015-2

Keywords

Search

Navigation