Regioselectivity of Hyoscyamine 6β-hydroxylase-catalysed Hydroxylation As Revealed by High-resolution Structural Information and QM/MM Calculations

Overview

Journal Dalton Trans

Publisher Royal Society of Chemistry

Specialty Chemistry

Date 2020 Mar 18

PMID 32182320

Citations 7

Authors

Anna Kluza

Zuzanna Wojdyla

Beata Mrugala

Katarzyna Kurpiewska

Przemyslaw J Porebski

Ewa Niedzialkowska

Wladek Minor

Manfred S Weiss

Tomasz Borowski

Affiliations

Soon will be listed here.

Abstract

Hyoscyamine 6β-hydroxylase (H6H) is a bifunctional non-heme 2-oxoglutarate/Fe2+-dependent dioxygenase that catalyzes the two final steps in the biosynthesis of scopolamine. Based on high resolution crystal structures of H6H from Datura metel, detailed information on substrate binding was obtained that provided insights into the onset of the enzymatic process. In particular, the role of two prominent residues was revealed - Glu-116 that interacts with the tertiary amine located on the hyoscyamine tropane moiety and Tyr-326 that forms CH-π hydrogen bonds with the hyoscyamine phenyl ring. The structures were used as the basis for QM/MM calculations that provided an explanation for the regioselectivity of the hydroxylation reaction on the hyoscyamine tropane moiety (C6 vs. C7) and quantified contributions of active site residues to respective barrier heights.

Citing Articles

Optimized Substrate Positioning Enables Switches in the C-H Cleavage Site and Reaction Outcome in the Hydroxylation-Epoxidation Sequence Catalyzed by Hyoscyamine 6β-Hydroxylase.

Wenger E, Martinie R, Ushimaru R, Pollock C, Sil D, Li A J Am Chem Soc. 2024; 146(35):24271-24287.

PMID: 39172701 PMC: 11374477. DOI: 10.1021/jacs.4c04406.

The Unique Role of the Second Coordination Sphere to Unlock and Control Catalysis in Nonheme Fe(II)/2-Oxoglutarate Histone Demethylase KDM2A.

Thomas M, Rifayee S, Chaturvedi S, Gorantla K, White W, Wildey J Inorg Chem. 2024; 63(23):10737-10755.

PMID: 38781256 PMC: 11168414. DOI: 10.1021/acs.inorgchem.4c01365.

Three-membered ring formation catalyzed by α-ketoglutarate-dependent nonheme iron enzymes.

Ushimaru R J Nat Med. 2023; 78(1):21-32.

PMID: 37980694 PMC: 10764440. DOI: 10.1007/s11418-023-01760-4.

Systematic analysis and expression of Gossypium 2ODD superfamily highlight the roles of GhLDOXs responding to alkali and other abiotic stress in cotton.

Jiang T, Cui A, Cui Y, Cui R, Han M, Zhang Y BMC Plant Biol. 2023; 23(1):124.

PMID: 36869319 PMC: 9985220. DOI: 10.1186/s12870-023-04133-x.

Biotechnological Approaches on Engineering Medicinal Tropane Alkaloid Production in Plants.

Gong H, He P, Lan X, Zeng L, Liao Z Front Plant Sci. 2022; 13:924413.

PMID: 35720595 PMC: 9201383. DOI: 10.3389/fpls.2022.924413.

References

Maier J, Martinez C, Kasavajhala K, Wickstrom L, Hauser K, Simmerling C . ff14SB: Improving the Accuracy of Protein Side Chain and Backbone Parameters from ff99SB. J Chem Theory Comput. 2015; 11(8):3696-713. PMC: 4821407. DOI: 10.1021/acs.jctc.5b00255. View

Rana S, Prasad Biswas J, Sen A, Clemancey M, Blondin G, Latour J . Selective C-H halogenation over hydroxylation by non-heme iron(iv)-oxo. Chem Sci. 2018; 9(40):7843-7858. PMC: 6194801. DOI: 10.1039/c8sc02053a. View

Rosenthal R, Vogeli B, Quade N, Capitani G, Kiefer P, Vorholt J . The use of ene adducts to study and engineer enoyl-thioester reductases. Nat Chem Biol. 2015; 11(6):398-400. DOI: 10.1038/nchembio.1794. View

Ye S, Geng C, Shaik S, Neese F . Electronic structure analysis of multistate reactivity in transition metal catalyzed reactions: the case of C-H bond activation by non-heme iron(IV)-oxo cores. Phys Chem Chem Phys. 2013; 15(21):8017-30. DOI: 10.1039/c3cp00080j. View

Roe D, Cheatham 3rd T . PTRAJ and CPPTRAJ: Software for Processing and Analysis of Molecular Dynamics Trajectory Data. J Chem Theory Comput. 2015; 9(7):3084-95. DOI: 10.1021/ct400341p. View

Lan X, Zeng J, Liu K, Zhang F, Bai G, Chen M . Comparison of two hyoscyamine 6β-hydroxylases in engineering scopolamine biosynthesis in root cultures of Scopolia lurida. Biochem Biophys Res Commun. 2018; 497(1):25-31. DOI: 10.1016/j.bbrc.2018.01.173. View

Zarate R, El Jaber-Vazdekis N, Medina B, Ravelo A . Tailoring tropane alkaloid accumulation in transgenic hairy roots of Atropa baetica by over-expressing the gene encoding hyoscyamine 6beta-hydroxylase. Biotechnol Lett. 2006; 28(16):1271-7. DOI: 10.1007/s10529-006-9085-8. View

Williams C, Headd J, Moriarty N, Prisant M, Videau L, Deis L . MolProbity: More and better reference data for improved all-atom structure validation. Protein Sci. 2017; 27(1):293-315. PMC: 5734394. DOI: 10.1002/pro.3330. View

Xia K, Liu X, Zhang Q, Qiang W, Guo J, Lan X . Promoting scopolamine biosynthesis in transgenic Atropa belladonna plants with pmt and h6h overexpression under field conditions. Plant Physiol Biochem. 2016; 106:46-53. DOI: 10.1016/j.plaphy.2016.04.034. View

10.

Krissinel E, Henrick K . Secondary-structure matching (SSM), a new tool for fast protein structure alignment in three dimensions. Acta Crystallogr D Biol Crystallogr. 2004; 60(Pt 12 Pt 1):2256-68. DOI: 10.1107/S0907444904026460. View

11.

Diederichs K . Some aspects of quantitative analysis and correction of radiation damage. Acta Crystallogr D Biol Crystallogr. 2005; 62(Pt 1):96-101. DOI: 10.1107/S0907444905031537. View

12.

Zucker F, Champ P, Merritt E . Validation of crystallographic models containing TLS or other descriptions of anisotropy. Acta Crystallogr D Biol Crystallogr. 2010; 66(Pt 8):889-900. PMC: 2917275. DOI: 10.1107/S0907444910020421. View

13.

Winn M, Ballard C, Cowtan K, Dodson E, Emsley P, Evans P . Overview of the CCP4 suite and current developments. Acta Crystallogr D Biol Crystallogr. 2011; 67(Pt 4):235-42. PMC: 3069738. DOI: 10.1107/S0907444910045749. View

14.

Kai G, Liu Y, Wang X, Yang S, Fu X, Luo X . Functional identification of hyoscyamine 6β-hydroxylase from Anisodus acutangulus and overproduction of scopolamine in genetically-engineered Escherichia coli. Biotechnol Lett. 2011; 33(7):1361-5. DOI: 10.1007/s10529-011-0575-y. View

15.

Slabinski L, Jaroszewski L, Rodrigues A, Rychlewski L, Wilson I, Lesley S . The challenge of protein structure determination--lessons from structural genomics. Protein Sci. 2007; 16(11):2472-82. PMC: 2211687. DOI: 10.1110/ps.073037907. View

16.

Timmins A, Saint-Andre M, de Visser S . Understanding How Prolyl-4-hydroxylase Structure Steers a Ferryl Oxidant toward Scission of a Strong C-H Bond. J Am Chem Soc. 2017; 139(29):9855-9866. DOI: 10.1021/jacs.7b02839. View

17.

Murshudov G, Skubak P, Lebedev A, Pannu N, Steiner R, Nicholls R . REFMAC5 for the refinement of macromolecular crystal structures. Acta Crystallogr D Biol Crystallogr. 2011; 67(Pt 4):355-67. PMC: 3069751. DOI: 10.1107/S0907444911001314. View

18.

Zhao K, Zeng J, Zhao T, Zhang H, Qiu F, Yang C . Enhancing Tropane Alkaloid Production Based on the Functional Identification of Tropine-Forming Reductase in , a Tibetan Medicinal Plant. Front Plant Sci. 2017; 8:1745. PMC: 5650612. DOI: 10.3389/fpls.2017.01745. View

19.

Schlesinger D, Davidovich Rikanati R, Volis S, Faigenboim A, Vendramin V, Cattonaro F . Alkaloid chemodiversity in Mandragora spp. is associated with loss-of-functionality of MoH6H, a hyoscyamine 6β-hydroxylase gene. Plant Sci. 2019; 283:301-310. DOI: 10.1016/j.plantsci.2019.03.013. View

20.

Grabowski M, Langner K, Cymborowski M, Porebski P, Sroka P, Zheng H . A public database of macromolecular diffraction experiments. Acta Crystallogr D Struct Biol. 2016; 72(Pt 11):1181-1193. PMC: 5108346. DOI: 10.1107/S2059798316014716. View