Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Special Feature: Review Article
  • Published:

Genome mining-driven discovery of enzymes catalyzing stereodivergent transformations

The Journal of Antibiotics volume 79, pages 183–194 (2026) Cite this article

Subjects

Abstract

Natural products provide privileged scaffolds for drug discovery, yet their stereochemical complexity often exceeds the limits of synthetic chemistry. Genome mining has emerged as a transformative strategy to uncover cryptic biosynthetic gene clusters and enzymes with noncanonical activities. Recent studies have revealed enzymes exhibiting unusual stereoselectivities, thereby expanding the enzymatic repertoire for constructing complex chiral architectures. Comparative analyses indicated that subtle variations in sequence and active-site environments produce diverse stereochemical outcomes across enzyme families. This review highlights representative examples of stereodivergent enzymes identified through genome- or sequence-guided approaches, emphasizing their substrate scope, catalytic mechanisms, and stereocontrol features. These advances not only deepen our mechanistic understanding of stereoselectivity but also lay the groundwork for rational enzyme engineering and the development of next-generation biocatalysts in pharmaceutical synthesis.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on SpringerLink
  • Instant access to the full article PDF.

USD 39.95

Prices may be subject to local taxes which are calculated during checkout

Fig. 1
The alternative text for this image may have been generated using AI.
Fig. 2
The alternative text for this image may have been generated using AI.
Fig. 3
The alternative text for this image may have been generated using AI.
Fig. 4
The alternative text for this image may have been generated using AI.
Fig. 5
The alternative text for this image may have been generated using AI.
Fig. 6
The alternative text for this image may have been generated using AI.
Fig. 7
The alternative text for this image may have been generated using AI.
Fig. 8
The alternative text for this image may have been generated using AI.
Fig. 9
The alternative text for this image may have been generated using AI.

Similar content being viewed by others

References

  1. Chopra B, Dhingra AK. Natural Products: A Lead for Drug Discovery and Development. Phytother Res. 2021;35:4660–702. https://doi.org/10.1002/ptr.7099.

    Article  PubMed  Google Scholar 

  2. Scherlach K, Hertweck C. Mining and Unearthing Hidden Biosynthetic Potential. Nat Commun. 2021;12:3864. https://doi.org/10.1038/s41467-021-24133-5.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. Galbiati A, Zana A, Borsari C, Persico M, Bova S, Tkachuk O, Corfu AI, Tamborini L, Basilico N, Fattorusso C, Bruno S, Parapini S, Conti P. Role of stereochemistry on the biological activity of nature-inspired 3-Br-acivicin isomers and derivatives. Molecules. 2023;28:3172 https://doi.org/10.3390/molecules28073172.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Scott KA, Ropek N, Melillo B, Schreiber SL, Cravatt BF, Vinogradova EV. Stereochemical Diversity as a Source of Discovery in Chemical Biology. Curr Res Chem Biol. 2022;2:100028. https://doi.org/10.1016/j.crchbi.2022.100028.

    Article  CAS  Google Scholar 

  5. Kawasaki T, Kuzuyama T, Kuwamori Y, Matsuura N, Itoh N, Furihata K, Seto H, Dairi T. Presence of Copalyl Diphosphate Synthase Gene in an Actinomycete Possessing the Mevalonate Pathway. J Antibiot. 2004;57:739–47. https://doi.org/10.7164/antibiotics.57.739.

    Article  CAS  Google Scholar 

  6. Ikeda H, Shin-ya K, Nagamitsu T, Tomoda H. Biosynthesis of Mercapturic Acid Derivative of the Labdane-Type Diterpene, Cyslabdan That Potentiates Imipenem Activity against Methicillin-Resistant Staphylococcus Aureus: Cyslabdan Is Generated by Mycothiol-Mediated Xenobiotic Detoxification. J Ind Microbiol Biotechnol. 2016;43:325–42. https://doi.org/10.1007/s10295-015-1694-6.

    Article  CAS  PubMed  Google Scholar 

  7. Yu J, Shiraishi T, Taizoumbe KA, Karasuno Y, Yoshida A, Nishiyama M, Dickschat JS, Kuzuyama T. Mechanistic Characterization of Diterpene Synthase Pairs for Tricyclic Diterpenes from Cyanobacteria. J Am Chem Soc. 2025;147:11896–905. https://doi.org/10.1021/jacs.4c16710.

    Article  CAS  PubMed  Google Scholar 

  8. Behshad E, Ruzicka FJ, Mansoorabadi SO, Chen D, Reed GH, Frey PA. Enantiomeric Free Radicals and Enzymatic Control of Stereochemistry in a Radical Mechanism:  The Case of Lysine 2,3-Aminomutases. Biochemistry. 2006;45:12639–46. https://doi.org/10.1021/bi061328t.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Ushimaru R, Cha L, Shimo S, Li X, Paris JC, Mori T, Miyamoto K, Coffer L, Uchiyama M, Guo Y, Chang W, Abe I. Mechanistic Analysis of Stereodivergent Nitroalkane Cyclopropanation Catalyzed by Nonheme Iron Enzymes. J Am Chem Soc. 2023;145:24210–7. https://doi.org/10.1021/jacs.3c08413.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Cha L, Paris JC, Zanella B, Spletzer M, Yao A, Guo Y, Chang W. Mechanistic Studies of Aziridine Formation Catalyzed by Mononuclear Non-Heme Iron Enzymes. J Am Chem Soc. 2023;145:6240–6. https://doi.org/10.1021/jacs.2c12664.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Ruzicka FJ, Lieder KW, Frey PA. Lysine 2,3-Aminomutase from Clostridium Subterminale SB4: Mass Spectral Characterization of Cyanogen Bromide-Treated Peptides and Cloning, Sequencing, and Expression of the Gene kamA in Escherichia Coli. J Bacteriol. 2000;182:469–76. https://doi.org/10.1128/jb.182.2.469-476.2000.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Ma C, Wang W, Zhang K, Zhang F, Chang Y, Sun C, Che Q, Zhu T, Zhang G, Li D. Exploring the Diverse Landscape of Fungal Cytochrome P450-Catalyzed Regio- and Stereoselective Dimerization of Diketopiperazines. Adv Sci. 2024;11:2310018. https://doi.org/10.1002/advs.202310018.

    Article  CAS  Google Scholar 

  13. Hara R, Kino K. Enzymatic Reactions and Microorganisms Producing the Various Isomers of Hydroxyproline. Appl Microbiol Biotechnol. 2020;104:4771–9. https://doi.org/10.1007/s00253-020-10603-1.

    Article  CAS  PubMed  Google Scholar 

  14. Mori H, Shibasaki T, Uozaki Y, Ochiai K, Ozaki A. Detection of Novel Proline 3-Hydroxylase Activities in Streptomyces and Bacillus Spp. by Regio- and Stereospecific Hydroxylation of l-Proline. Appl Environ Microbiol. 1996;62:1903–7. https://doi.org/10.1128/aem.62.6.1903-1907.1996.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Shibasaki T, Mori H, Chiba S, Ozaki A. Microbial Proline 4-Hydroxylase Screening and Gene Cloning. Appl Environ Microbiol. 1999;65:4028–31. https://doi.org/10.1128/AEM.65.9.4028-4031.1999.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Islam MS, Leissing TM, Chowdhury R, Hopkinson RJ, Schofield CJ. 2-Oxoglutarate-Dependent Oxygenases. Annu Rev Biochem. 2018;87:585–620. https://doi.org/10.1146/annurev-biochem-061516-044724.

    Article  CAS  PubMed  Google Scholar 

  17. Martinez S, Hausinger RP. Catalytic Mechanisms of Fe(II)- and 2-Oxoglutarate-Dependent Oxygenases. J Biol Chem. 2015;290:20702–11. https://doi.org/10.1074/jbc.R115.648691.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Hausinger, RP Biochemical Diversity of 2-Oxoglutarate-Dependent Oxygenases. In 2-Oxoglutarate-Dependent Oxygenases; Schofield, C, Hausinger, R, Eds.; The Royal Society of Chemistry, 2015, 1–58. https://doi.org/10.1039/9781782621959-00001.

  19. Krebs C, Galonić Fujimori D, Walsh CT, Bollinger JM Jr. Non-Heme Fe(IV)–Oxo Intermediates. Acc Chem Res. 2007;40:484–92. https://doi.org/10.1021/ar700066p.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Ushimaru R, Abe I. Unusual Dioxygen-Dependent Reactions Catalyzed by Nonheme Iron Enzymes in Natural Product Biosynthesis. ACS Catal. 2023;13:1045–76. https://doi.org/10.1021/acscatal.2c05247.

    Article  CAS  Google Scholar 

  21. Zhao, S; Wu, L; Xu, Y; Nie, Y Fe (II) and 2-Oxoglutarate-Dependent Dioxygenases for Natural Product Synthesis: Molecular Insights into Reaction Diversity. 2025, 42, 67–92. https://doi.org/10.1039/D4NP00030G.

  22. Jing X, Wang X, Zhang W, An J, Luo P, Nie Y, Xu Y. Highly Regioselective and Stereoselective Hydroxylation of Free Amino Acids by a 2-Oxoglutarate-Dependent Dioxygenase from Kutzneria Albida. ACS Omega. 2019;4:8350–8. https://doi.org/10.1021/acsomega.9b00983.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Hara R, Kino K. Characterization of Novel 2-Oxoglutarate Dependent Dioxygenases Converting l-Proline to Cis-4-Hydroxy-l-Proline. Biochem Biophys Res Commun. 2009;379:882–6. https://doi.org/10.1016/j.bbrc.2008.12.158.

    Article  CAS  PubMed  Google Scholar 

  24. Koehntop KD, Emerson JP, Que L. The 2-His-1-Carboxylate Facial Triad: A Versatile Platform for Dioxygen Activation by Mononuclear Non-Heme Iron(II) Enzymes. JBIC J Biol Inorg Chem. 2005;10:87–93. https://doi.org/10.1007/s00775-005-0624-x.

    Article  CAS  PubMed  Google Scholar 

  25. Hara R, Uchiumi N, Kino K. Identification and Characterization of 2-Oxoglutarate-Dependent Dioxygenases Catalyzing Selective Cis-Hydroxylation of Proline and Pipecolinic Acid from Actinomycetes. J Biotechnol. 2014;172:55–58. https://doi.org/10.1016/j.jbiotec.2013.12.003.

    Article  CAS  PubMed  Google Scholar 

  26. Hara R, Uchiumi N, Okamoto N, Kino K. Regio- and Stereoselective Oxygenation of Proline Derivatives by Using Microbial 2-Oxoglutarate-Dependent Dioxygenases. Biosci Biotechnol Biochem. 2014;78:1384–8. https://doi.org/10.1080/09168451.2014.918490.

    Article  CAS  PubMed  Google Scholar 

  27. Houwaart S, Youssar L, Hüttel W. Pneumocandin Biosynthesis: Involvement of a Trans-Selective Proline Hydroxylase. ChemBioChem. 2014;15:2365–9. https://doi.org/10.1002/cbic.201402175.

    Article  CAS  PubMed  Google Scholar 

  28. Mattay J, Houwaart S, Hüttel W. Cryptic Production of Trans-3-Hydroxyproline in Echinocandin B Biosynthesis. Appl Environ Microbiol. 2018;84:e02370–17. https://doi.org/10.1128/AEM.02370-17.

    Article  PubMed  PubMed Central  Google Scholar 

  29. Kawai S, Sugaya Y, Hagihara R, Tomita H, Katsuyama Y, Ohnishi Y. Complete Biosynthetic Pathway of Alazopeptin, a Tripeptide Consisting of Two Molecules of 6-Diazo-5-Oxo-l-Norleucine and One Molecule of Alanine. Angew Chem Int Ed. 2021;60:10319–25. https://doi.org/10.1002/anie.202100462.

    Article  CAS  Google Scholar 

  30. Kawai S, Moriga K, Nirdnoy W, Hara R, Ogawa J, Katsuyama Y, Ohnishi Y. Identification of Two Distinct Stereoselective Lysine 5-Hydroxylases by Genome Mining Based on Alazopeptin Biosynthetic Enzymes. Chem Eur J. 2025;31:e202404790. https://doi.org/10.1002/chem.202404790.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Shimo S, Ushimaru R, Engelbrecht A, Harada M, Miyamoto K, Kulik A, Uchiyama M, Kaysser L, Abe I. Stereodivergent Nitrocyclopropane Formation during Biosynthesis of Belactosins and Hormaomycins. J Am Chem Soc. 2021;143:18413–8. https://doi.org/10.1021/jacs.1c10201.

    Article  CAS  PubMed  Google Scholar 

  32. Li X, Shimaya R, Dairi T, Chang W, Ogasawara Y. Identification of Cyclopropane Formation in the Biosyntheses of Hormaomycins and Belactosins: Sequential Nitration and Cyclopropanation by Metalloenzymes. Angew Chem Int Ed. 2022;61:e202113189. https://doi.org/10.1002/anie.202113189.

    Article  CAS  Google Scholar 

  33. Skirboll SS, Gangopadhyay M, Phan HN, Hartsell J, Mudireddy A, Hilovsky D, Swartz PD, Liu X, Guo Y, Makris TM. The Heme Oxygenase-Like Diiron Enzyme HrmI Reveals Altered Regulatory Mechanisms for Dioxygen Activation and Substrate N-Oxygenation. J Am Chem Soc. 2025;147:30210–21. https://doi.org/10.1021/jacs.5c08814.

    Article  CAS  PubMed  Google Scholar 

  34. Zheng Y, Li X, Cha L, Paris JC, Michael C, Ushimaru R, Ogasawara Y, Abe I, Guo Y, Chang W. Comparison of a Nonheme Iron Cyclopropanase with a Homologous Hydroxylase Reveals Mechanistic Features Associated with Distinct Reaction Outcomes. J Am Chem Soc. 2025;147:6162–70. https://doi.org/10.1021/jacs.4c17741.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Ushimaru R. Three-Membered Ring Formation Catalyzed by α-Ketoglutarate-Dependent Nonheme Iron Enzymes. J Nat Med. 2024;78:21–32. https://doi.org/10.1007/s11418-023-01760-4.

    Article  CAS  PubMed  Google Scholar 

  36. Gao X, Chooi Y, Ames BD, Wang P, Walsh CT, Tang Y. Fungal Indole Alkaloid Biosynthesis: Genetic and Biochemical Investigation of the Tryptoquialanine Pathway in Penicillium Aethiopicum. J Am Chem Soc. 2011;133:2729–41. https://doi.org/10.1021/ja1101085.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  37. Bunno R, Awakawa T, Mori T, Abe I. Aziridine Formation by a FeII/α-Ketoglutarate Dependent Oxygenase and 2-Aminoisobutyrate Biosynthesis in Fungi. Angew Chem Int Ed. 2021;60:15827–31. https://doi.org/10.1002/anie.202104644.

    Article  CAS  Google Scholar 

  38. Tao H, Ushimaru R, Awakawa T, Mori T, Uchiyama M, Abe I. Stereoselectivity and Substrate Specificity of the Fe(II)/α-Ketoglutarate-Dependent Oxygenase TqaL. J Am Chem Soc. 2022;144:21512–20. https://doi.org/10.1021/jacs.2c08116.

    Article  CAS  PubMed  Google Scholar 

  39. Zhou L, Awakawa T, Ushimaru R, Kanaida M, Abe I. Characterization of Aziridine-Forming α-Ketoglutarate-Dependent Oxygenase in l-Isovaline Biosynthesis. Org Lett. 2024;26:724–7. https://doi.org/10.1021/acs.orglett.3c04185.

    Article  CAS  PubMed  Google Scholar 

  40. Chirpich TP, Zappia V, Costilow RN, Barker HA. Lysine 2,3-Aminomutase: Purification and Properties of a Pyridoxal Phosphate and S-Adenosylmethionine-Activated Enzyme. J Biol Chem. 1970;245:1778–89. https://doi.org/10.1016/S0021-9258(19)77160-9.

    Article  CAS  PubMed  Google Scholar 

  41. Stadtman, TC Lysine Metabolism by Clostridia. In Advances in Enzymology and Related Areas of Molecular Biology; John Wiley & Sons, Ltd, 1973; 413-48. https://doi.org/10.1002/9780470122839.ch9.

  42. Aberhart DJ, Gould SJ, Lin HJ, Thiruvengadam TK, Weiller BH. Stereochemistry of Lysine 2,3-Aminomutase Isolated from Clostridium Subterminale Strain SB4. J Am Chem Soc. 1983;105:5461–70. https://doi.org/10.1021/ja00354a046.

    Article  CAS  Google Scholar 

  43. Lepore BW, Ruzicka FJ, Frey PA, Ringe D. The X-Ray Crystal Structure of Lysine-2,3-Aminomutase from Clostridium Subterminale. Proc Natl Acad Sci. 2005;102:13819–24. https://doi.org/10.1073/pnas.0505726102.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  44. Frey PA. Radical Mechanisms of Enzymatic Catalysis1. Annu Rev Biochem. 2001;70:121–48. https://doi.org/10.1146/annurev.biochem.70.1.121.

    Article  CAS  PubMed  Google Scholar 

  45. Frey PA. Transient Intermediates in Enzymology, 1964–2008. J Biol Chem. 2015;290:10610–26. https://doi.org/10.1074/jbc.X115.650879.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  46. Zhao J, Ji W, Ji X, Zhang Q. Biochemical Characterization of an Arginine 2,3-Aminomutase with Dual Substrate Specificity. Chin J Chem. 2020;38:959–62. https://doi.org/10.1002/cjoc.202000119.

    Article  CAS  Google Scholar 

  47. Ushimaru R, Abe I. C–N and C–S Bond Formation by Cytochrome P450 Enzymes. Trends Chem. 2023;5:526–36. https://doi.org/10.1016/j.trechm.2023.04.008.

    Article  CAS  Google Scholar 

  48. Brooks BW, van den Berg S, Dreier DA, LaLone CA, Owen SF, Raimondo S, Zhang X. Towards Precision Ecotoxicology: Leveraging Evolutionary Conservation of Pharmaceutical and Personal Care Product Targets to Understand Adverse Outcomes Across Species and Life Stages. Environ Toxicol Chem. 2024;43:526–36. https://doi.org/10.1002/etc.5754.

    Article  CAS  PubMed  Google Scholar 

  49. Saruwatari T, Yagishita F, Mino T, Noguchi H, Hotta K, Watanabe K. Cytochrome P450 as Dimerization Catalyst in Diketopiperazine Alkaloid Biosynthesis. ChemBioChem. 2014;15:656–9. https://doi.org/10.1002/cbic.201300751.

    Article  CAS  PubMed  Google Scholar 

  50. Tian W, Sun C, Zheng M, Harmer JR, Yu M, Zhang Y, Peng H, Zhu D, Deng Z, Chen S-L, Mobli M, Jia X, Qu X. Efficient Biosynthesis of Heterodimeric C3-Aryl Pyrroloindoline Alkaloids. Nat Commun. 2018;9:4428. https://doi.org/10.1038/s41467-018-06528-z.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  51. Shende VV, Khatri Y, Newmister SA, Sanders JN, Lindovska P, Yu F, Doyon TJ, Kim J, Houk KN, Movassaghi M, Sherman DH. Structure and Function of NzeB, a Versatile C–C and C–N Bond-Forming Diketopiperazine Dimerase. J Am Chem Soc. 2020;142:17413–24. https://doi.org/10.1021/jacs.0c06312.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  52. Shibasaki T, Mori H, Ozaki A. Enzymatic Production of Trans-4-Hydroxy-L-Proline by Regio- and Stereospecific Hydroxylation of L-Proline. Biosci Biotechnol Biochem. 2000;64:746–50. https://doi.org/10.1271/bbb.64.746.

    Article  CAS  PubMed  Google Scholar 

  53. Wang X, Liu J, Zhao J, Ni X, Zheng P, Guo X, Sun C, Sun J, Ma Y. Efficient Production of Trans-4-Hydroxy-l-Proline from Glucose Using a New Trans-Proline 4-Hydroxylase in Escherichia Coli. J Biosci Bioeng. 2018;126:470–7. https://doi.org/10.1016/j.jbiosc.2018.04.012.

    Article  CAS  PubMed  Google Scholar 

Download references

Funding

This work was supported in part by Grants-in-Aid for Scientific Research from the Ministry of Education, Culture, Sports, Science and Technology, Japan (Japan Society for the Promotion of Science KAKENHI Grant Numbers JP22H05123, JP24H01309, JP25H02006, JP25K02417) and Japan Science and Technology Agency (FOREST Grant Number JPMJFR2305).

Author information

Authors and Affiliations

  1. Institute for Advanced Study and Department of Chemistry, Graduate School of Science, Kyushu University, 744 Motooka, Nishi-ku, Fukuoka, 819-0395, Japan

    Jiayi Yu & Richiro Ushimaru

  2. FOREST, Japan Science and Technology Agency, 4-1-8 Honcho, Kawaguchi, Saitama, 332-0012, Japan

    Richiro Ushimaru

Authors
  1. Jiayi Yu
    View author publications

    Search author on:PubMed Google Scholar

  2. Richiro Ushimaru
    View author publications

    Search author on:PubMed Google Scholar

Corresponding author

Correspondence to Richiro Ushimaru.

Ethics declarations

Conflict of interest

The authors declare no conflict of interest.

Additional information

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

Rights and permissions

Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Yu, J., Ushimaru, R. Genome mining-driven discovery of enzymes catalyzing stereodivergent transformations. J Antibiot 79, 183–194 (2026). https://doi.org/10.1038/s41429-025-00881-0

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Version of record:

  • Issue date:

  • DOI: https://doi.org/10.1038/s41429-025-00881-0

Search

Advanced search

Quick links