Structure and function of a flavin-dependent S-monooxygenase from garlic (Allium sativum)

Valentino, Hannah; Campbell, Ashley C.; Schuermann, Jonathan P.; Sultana, Nazneen; Nam, Han G.; LeBlanc, Sophie; Tanner, John J.; Sobrado, Pablo

doi:10.1074/jbc.ra120.014484

Structure and function of a flavin-dependent S-monooxygenase from garlic (Allium sativum)

Journal Article · Thu Jun 11 00:00:00 EDT 2020 · Journal of Biological Chemistry

DOI:https://doi.org/10.1074/jbc.ra120.014484· OSTI ID:1647382

Valentino, Hannah ^[1]; Campbell, Ashley C. ^[2]; Schuermann, Jonathan P. ^[3]; Sultana, Nazneen ^[1]; Nam, Han G. ^[1]; ^[1]; ^[2]; ^[1]

Virginia Polytechnic Inst. and State Univ. (Virginia Tech), Blacksburg, VA (United States)
Univ. of Missouri, Columbia, MO (United States)
Cornell Univ., Ithaca, NY (United States)

Allicin is a component of the characteristic smell and flavor of garlic (Allium sativum). A flavin-containing monooxygenase (FMO) produced by A. sativum (AsFMO) was previously proposed to oxidize S-allyl-L-cysteine (SAC) to alliin, an allicin precursor. In this paper, we present a kinetic and structural characterization of AsFMO that suggests a possible contradiction to this proposal. Results of steady-state kinetic analyses revealed that AsFMO exhibited negligible activity with SAC; however, the enzyme was highly active with L-cysteine, N-acetyl-L-cysteine, and allyl mercaptan. We found that allyl mercaptan with NADPH was the preferred substrate-cofactor combination. Rapid-reaction kinetic analyses showed that NADPH binds tightly ($$K_D$$ of ~2 μM) to AsFMO and that the hydride transfer occurs with pro-R stereospecificity. We detected the formation of a long-wavelength band when AsFMO was reduced by NADPH, probably representing the formation of a charge-transfer complex. In the absence of substrate, the reduced enzyme, in complex with NADP⁺, reacted with oxygen and formed an intermediate with a spectrum characteristic of C4a-hydroperoxyflavin, which decays several orders of magnitude more slowly than the $$k$$_cat. The presence of substrate enhanced C4a-hydroperoxyflavin formation and, upon hydroxylation, oxidation occurred with a rate constant similar to the $$k$$_cat. The structure of AsFMO complexed with FAD at 2.08-Å resolution features two domains for binding of FAD and NADPH, representative of class B flavin monooxygenases. These biochemical and structural results are consistent with AsFMO being an S-monooxygenase involved in allicin biosynthesis through direct formation of sulfenic acid and not SAC oxidation.

View Accepted Manuscript (DOE)

Research Organization:: Argonne National Laboratory (ANL), Argonne, IL (United States). Advanced Photon Source (APS)

Sponsoring Organization:: National Institute of General Medical Sciences (NIGMS); National Institutes of Health (NIH); USDOE Office of Science (SC)

Grant/Contract Number:: AC02-06CH11357

OSTI ID:: 1647382

Journal Information:: Journal of Biological Chemistry, Journal Name: Journal of Biological Chemistry Journal Issue: 32 Vol. 295; ISSN 0021-9258

Publisher:: American Society for Biochemistry and Molecular BiologyCopyright Statement

Country of Publication:: United States

Language:: ENGLISH

References (61)

Garlic-derived anticancer agents: Structure and biological activity of ajoene Kaschula, Catherine H.; Hunter, Roger; Parker, M. Iqbal BioFactors https://doi.org/10.1002/biof.76	journal	January 2010
Automated Structure Solution with the PHENIX Suite Zwart, Peter H.; Afonine, Pavel V.; Grosse-Kunstleve, Ralf W. Methods in Molecular Biology https://doi.org/10.1007/978-1-60327-058-8_28	book	January 2008
Antimicrobial properties of Allium sativum (garlic) Harris, J.; S., Cottrell; S., Plummer Applied Microbiology and Biotechnology, Vol. 57, Issue 3 https://doi.org/10.1007/s002530100722	journal	October 2001
Volatile distribution in garlic (Allium sativum L.) by solid phase microextraction (SPME) with different processing conditions Kim, Na Young; Park, Min Hee; Jang, Eun Yeong Food Science and Biotechnology, Vol. 20, Issue 3 https://doi.org/10.1007/s10068-011-0108-4	journal	June 2011
Allium ursinum: botanical, phytochemical and pharmacological overview Sobolewska, Danuta; Podolak, Irma; Makowska-Wąs, Justyna Phytochemistry Reviews, Vol. 14, Issue 1 https://doi.org/10.1007/s11101-013-9334-0	journal	December 2013
Chemical applications of Class B flavoprotein monooxygenases Catucci, Gianluca; Gao, Chongliang; Sadeghi, Sheila J. Rendiconti Lincei, Vol. 28, Issue S1 https://doi.org/10.1007/s12210-016-0583-x	journal	November 2016
Solvent content of protein crystals Matthews, B. W. Journal of Molecular Biology, Vol. 33, Issue 2 https://doi.org/10.1016/0022-2836(68)90205-2	journal	April 1968
Control of catalysis in flavin-dependent monooxygenases Palfey, Bruce A.; McDonald, Claudia A. Archives of Biochemistry and Biophysics, Vol. 493, Issue 1 https://doi.org/10.1016/j.abb.2009.11.028	journal	January 2010
Flavin dependent monooxygenases Huijbers, Mieke M. E.; Montersino, Stefania; Westphal, Adrie H. Archives of Biochemistry and Biophysics, Vol. 544 https://doi.org/10.1016/j.abb.2013.12.005	journal	February 2014
Mechanistic studies on the flavin-dependent N6-lysine monooxygenase MbsG reveal an unusual control for catalysis Robinson, Reeder M.; Rodriguez, Pedro J.; Sobrado, Pablo Archives of Biochemistry and Biophysics, Vol. 550-551 https://doi.org/10.1016/j.abb.2014.04.006	journal	May 2014
Dual role of NADP(H) in the reaction of a flavin dependent N-hydroxylating monooxygenase Romero, Elvira; Fedkenheuer, Michael; Chocklett, Samuel W. Biochimica et Biophysica Acta (BBA) - Proteins and Proteomics, Vol. 1824, Issue 6 https://doi.org/10.1016/j.bbapap.2012.03.004	journal	June 2012
Mammalian flavin-containing monooxygenase (FMO) as a source of hydrogen peroxide Siddens, Lisbeth K.; Krueger, Sharon K.; Henderson, Marilyn C. Biochemical Pharmacology, Vol. 89, Issue 1 https://doi.org/10.1016/j.bcp.2014.02.006	journal	May 2014
Flavin Monooxygenase-Generated N-Hydroxypipecolic Acid Is a Critical Element of Plant Systemic Immunity Hartmann, Michael; Zeier, Tatyana; Bernsdorff, Friederike Cell, Vol. 173, Issue 2 https://doi.org/10.1016/j.cell.2018.02.049	journal	April 2018
The analysis of onion and garlic Lanzotti, Virginia Journal of Chromatography A, Vol. 1112, Issue 1-2 https://doi.org/10.1016/j.chroma.2005.12.016	journal	April 2006
Flavoprotein monooxygenases, a diverse class of oxidative biocatalysts van Berkel, W. J. H.; Kamerbeek, N. M.; Fraaije, M. W. Journal of Biotechnology, Vol. 124, Issue 4 https://doi.org/10.1016/j.jbiotec.2006.03.044	journal	August 2006
Inference of Macromolecular Assemblies from Crystalline State Krissinel, Evgeny; Henrick, Kim Journal of Molecular Biology, Vol. 372, Issue 3 https://doi.org/10.1016/j.jmb.2007.05.022	journal	September 2007
Structural and functional analysis of bacterial flavin-containing monooxygenase reveals its ping-pong-type reaction mechanism Cho, Hyo Je; Cho, Ha Yeon; Kim, Kyung Jin Journal of Structural Biology, Vol. 175, Issue 1 https://doi.org/10.1016/j.jsb.2011.04.007	journal	July 2011
Headspace−GC–MS volatile profile of black garlic vs fresh garlic: Evolution along fermentation and behavior under heating Molina-Calle, María; Priego-Capote, Feliciano; Luque de Castro, María D. LWT, Vol. 80 https://doi.org/10.1016/j.lwt.2017.02.010	journal	July 2017
Validation of Structures in the Protein Data Bank Gore, Swanand; Sanz García, Eduardo; Hendrickx, Pieter M. S. Structure, Vol. 25, Issue 12 https://doi.org/10.1016/j.str.2017.10.009	journal	December 2017
Flavin-containing monooxygenases in plants: looking beyond detox Schlaich, Nikolaus L. Trends in Plant Science, Vol. 12, Issue 9 https://doi.org/10.1016/j.tplants.2007.08.009	journal	September 2007
Enzyme Kinetics of Different Types of Flavin-Dependent Monooxygenases Determine the Observable Contaminant Stable Isotope Fractionation Wijker, Reto S.; Pati, Sarah G.; Zeyer, Josef Environmental Science & Technology Letters, Vol. 2, Issue 11 https://doi.org/10.1021/acs.estlett.5b00254	journal	October 2015
Trapping Reactions of the Sulfenyl and Sulfinyl Tautomers of Sulfenic Acids Kumar, Murugaeson R.; Farmer, Patrick J. ACS Chemical Biology, Vol. 12, Issue 2 https://doi.org/10.1021/acschembio.6b00980	journal	December 2016
Norbornene Probes for the Detection of Cysteine Sulfenic Acid in Cells Alcock, Lisa J.; Oliveira, Bruno L.; Deery, Michael J. ACS Chemical Biology, Vol. 14, Issue 4 https://doi.org/10.1021/acschembio.8b01104	journal	March 2019
Aspergillus fumigatus SidA Is a Highly Specific Ornithine Hydroxylase with Bound Flavin Cofactor Chocklett, Samuel W.; Sobrado, Pablo Biochemistry, Vol. 49, Issue 31 https://doi.org/10.1021/bi100291n	journal	August 2010
Structural Insight into the Mechanism of Oxygen Activation and Substrate Selectivity of Flavin-Dependent N-Hydroxylating Monooxygenases Franceschini, Stefano; Fedkenheuer, Michael; Vogelaar, Nancy J. Biochemistry, Vol. 51, Issue 36 https://doi.org/10.1021/bi301072w	journal	August 2012
C4a-Hydroperoxyflavin Formation in N -Hydroxylating Flavin Monooxygenases Is Mediated by the 2′-OH of the Nicotinamide Ribose of NADP ⁺ Robinson, Reeder; Badieyan, Somayesadat; Sobrado, Pablo Biochemistry, Vol. 52, Issue 51 https://doi.org/10.1021/bi4014903	journal	November 2013
Oxidation of cysteine to cystine using hydrogen peroxide Hill, John W.; Coy, Robert B.; Lewandowski, Peter E. Journal of Chemical Education, Vol. 67, Issue 2 https://doi.org/10.1021/ed067p172	journal	February 1990
Identification of the NAD(P)H Binding Site of Eukaryotic UDP-Galactopyranose Mutase Dhatwalia, Richa; Singh, Harkewal; Solano, Luis M. Journal of the American Chemical Society, Vol. 134, Issue 43 https://doi.org/10.1021/ja308188z	journal	October 2012
Unusual Cystine Lyase Activity of the Enzyme Alliinase: Direct Formation of Polysulphides Keusgen, Michael Planta Medica, Vol. 74, Issue 1 https://doi.org/10.1055/s-2007-993777	journal	January 2008
Mechanism of action of a flavin-containing monooxygenase Eswaramoorthy, S.; Bonanno, J. B.; Burley, S. K. Proceedings of the National Academy of Sciences, Vol. 103, Issue 26 https://doi.org/10.1073/pnas.0602398103	journal	June 2006
Revealing the moonlighting role of NADP in the structure of a flavin-containing monooxygenase Alfieri, A.; Malito, E.; Orru, R. Proceedings of the National Academy of Sciences, Vol. 105, Issue 18 https://doi.org/10.1073/pnas.0800859105	journal	April 2008
Crystal Structures and Small-angle X-ray Scattering Analysis of UDP-galactopyranose Mutase from the Pathogenic Fungus Aspergillus fumigatus Dhatwalia, Richa; Singh, Harkewal; Oppenheimer, Michelle Journal of Biological Chemistry, Vol. 287, Issue 12 https://doi.org/10.1074/jbc.M111.327536	journal	January 2012
The Biochemical Mechanism of Auxin Biosynthesis by an Arabidopsis YUCCA Flavin-containing Monooxygenase Dai, Xinhua; Mashiguchi, Kiyoshi; Chen, Qingguo Journal of Biological Chemistry, Vol. 288, Issue 3 https://doi.org/10.1074/jbc.M112.424077	journal	November 2012
An Unprecedented NADPH Domain Conformation in Lysine Monooxygenase NbtG Provides Insights into Uncoupling of Oxygen Consumption from Substrate Hydroxylation Binda, Claudia; Robinson, Reeder M.; Martin del Campo, Julia S. Journal of Biological Chemistry, Vol. 290, Issue 20 https://doi.org/10.1074/jbc.M114.629485	journal	March 2015
High-Performance Liquid Chromatographic Analysis of Free Amino Acids in Fruit Juices Using Derivatization with 9-Fluorenylmethyl-Chloroformate Fabiani, A.; Versari, A.; Parpinello, G. P. Journal of Chromatographic Science, Vol. 40, Issue 1 https://doi.org/10.1093/chromsci/40.1.14	journal	January 2002
Biosynthesis of the flavour precursors of onion and garlic Jones, M. G. Journal of Experimental Botany, Vol. 55, Issue 404 https://doi.org/10.1093/jxb/erh138	journal	July 2004
S-Alk(en)ylcysteine sulfoxides in the genus Allium: proposed biosynthesis, chemical conversion, and bioactivities Yoshimoto, Naoko; Saito, Kazuki Journal of Experimental Botany, Vol. 70, Issue 16 https://doi.org/10.1093/jxb/erz243	journal	May 2019
Subclade of Flavin-Monooxygenases Involved in Aliphatic Glucosinolate Biosynthesis Li, Jing; Hansen, Bjarne Gram; Ober, James A. Plant Physiology, Vol. 148, Issue 3 https://doi.org/10.1104/pp.108.125757	journal	September 2008
Substructure solution with SHELXD Schneider, Thomas R.; Sheldrick, George M. Acta Crystallographica Section D Biological Crystallography, Vol. 58, Issue 10 https://doi.org/10.1107/S0907444902011678	journal	September 2002
Secondary-structure matching (SSM), a new tool for fast protein structure alignment in three dimensions Krissinel, E.; Henrick, K. Acta Crystallographica Section D Biological Crystallography, Vol. 60, Issue 12 https://doi.org/10.1107/S0907444904026460	journal	November 2004
MolProbity : all-atom structure validation for macromolecular crystallography Chen, Vincent B.; Arendall, W. Bryan; Headd, Jeffrey J. Acta Crystallographica Section D Biological Crystallography, Vol. 66, Issue 1 https://doi.org/10.1107/S0907444909042073	journal	December 2009
XDS Kabsch, Wolfgang Acta Crystallographica Section D Biological Crystallography, Vol. 66, Issue 2 https://doi.org/10.1107/S0907444909047337	journal	January 2010
Features and development of Coot Emsley, P.; Lohkamp, B.; Scott, W. G. Acta Crystallographica Section D Biological Crystallography, Vol. 66, Issue 4 https://doi.org/10.1107/S0907444910007493	journal	March 2010
Towards automated crystallographic structure refinement with phenix.refine Afonine, Pavel V.; Grosse-Kunstleve, Ralf W.; Echols, Nathaniel Acta Crystallographica Section D Biological Crystallography, Vol. 68, Issue 4 https://doi.org/10.1107/S0907444912001308	journal	March 2012
How good are my data and what is the resolution? Evans, Philip R.; Murshudov, Garib N. Acta Crystallographica Section D Biological Crystallography, Vol. 69, Issue 7 https://doi.org/10.1107/S0907444913000061	journal	June 2013
Polder maps: improving OMIT maps by excluding bulk solvent Liebschner, Dorothee; Afonine, Pavel V.; Moriarty, Nigel W. Acta Crystallographica Section D Structural Biology, Vol. 73, Issue 2, p. 148-157 https://doi.org/10.1107/S2059798316018210	journal	February 2017
Crystal structure of pyrrolizidine alkaloid N -oxygenase from the grasshopper Zonocerus variegatus Kubitza, Christian; Faust, Annette; Gutt, Miriam Acta Crystallographica Section D Structural Biology, Vol. 74, Issue 5 https://doi.org/10.1107/S2059798318003510	journal	April 2018
Identification of a flavin-monooxygenase as the S-oxygenating enzyme in aliphatic glucosinolate biosynthesis in Arabidopsis: Identification of a flavin monooxygenase Hansen, Bjarne G.; Kliebenstein, Daniel J.; Halkier, Barbara A. The Plant Journal, Vol. 50, Issue 5 https://doi.org/10.1111/j.1365-313X.2007.03101.x	journal	April 2007
Structural mechanism for bacterial oxidation of oceanic trimethylamine into trimethylamine N -oxide: NADP ⁺ bending is important for TMA oxidation Li, Chun-Yang; Chen, Xiu-Lan; Zhang, Dian Molecular Microbiology, Vol. 103, Issue 6 https://doi.org/10.1111/mmi.13605	journal	January 2017
Identification of a flavin-containing S -oxygenating monooxygenase involved in alliin biosynthesis in garlic Yoshimoto, Naoko; Onuma, Misato; Mizuno, Shinya The Plant Journal, Vol. 83, Issue 6 https://doi.org/10.1111/tpj.12954	journal	September 2015
Quantification and Characterization of Allicin in Garlic Extract Mansor, Nurlidia; Herng, Ho Jian; Samsudin, Sity Juaeiriah Journal of Medical and Bioengineering, Vol. 5, Issue 1 https://doi.org/10.12720/jomb.5.1.24-27	journal	January 2016
Organosulfur chemistry of garlic and onion: Recent results Block, Eric; Naganathan, S.; Putman, D. Pure and Applied Chemistry, Vol. 65, Issue 4 https://doi.org/10.1351/pac199365040625	journal	January 1993
Mechanism of Rifampicin Inactivation in Nocardia farcinica Abdelwahab, Heba; Martin Del Campo, Julia S.; Dai, Yumin PLOS ONE, Vol. 11, Issue 10 https://doi.org/10.1371/journal.pone.0162578	journal	October 2016
Two Novel Flavin-Containing Monooxygenases Involved in Biosynthesis of Aliphatic Glucosinolates Kong, Wenwen; Li, Jing; Yu, Qingyue Frontiers in Plant Science, Vol. 7 https://doi.org/10.3389/fpls.2016.01292	journal	August 2016
The “Green” FMOs: Diversity, Functionality and Application of Plant Flavoproteins Thodberg, Sara; Jakobsen Neilson, Elizabeth H. Catalysts, Vol. 10, Issue 3 https://doi.org/10.3390/catal10030329	journal	March 2020
Flavin-Dependent Enzymes in Cancer Prevention Wojcieszyńska, Danuta; Hupert-Kocurek, Katarzyna; Guzik, Urszula International Journal of Molecular Sciences, Vol. 13, Issue 12 https://doi.org/10.3390/ijms131216751	journal	December 2012
Allicin: Chemistry and Biological Properties Borlinghaus, Jan; Albrecht, Frank; Gruhlke, Martin Molecules, Vol. 19, Issue 8 https://doi.org/10.3390/molecules190812591	journal	August 2014
Selenium and Sulfur to Produce Allium Functional Crops González-Morales, Susana; Pérez-Labrada, Fabián; García-Enciso, Ema Molecules, Vol. 22, Issue 4 https://doi.org/10.3390/molecules22040558	journal	March 2017
Characteristics, biosynthesis, decomposition, metabolism and functions of the garlic odour precursor, S‑allyl‑l‑cysteine sulfoxide (Review) Yamaguchi, Yusuke; Kumagai, Hitomi Experimental and Therapeutic Medicine https://doi.org/10.3892/etm.2019.8385	journal	December 2019
Antioxidant activity and leukemia initiation prevention in�vitro and in�vivo by N‑acetyl‑L‑cysteine Shih, Wen‑Ling; Chang, Ching‑Dong; Chen, Hsiu‑Tsu Oncology Letters https://doi.org/10.3892/ol.2018.8864	journal	May 2018
Extracts from the history and medical properties of garlic Petrovska, BiljanaBauer; Cekovska, Svetlana Pharmacognosy Reviews, Vol. 4, Issue 7 https://doi.org/10.4103/0973-7847.65321	journal	January 2010

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Related Subjects

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CHEMISTRY
59 BASIC BIOLOGICAL SCIENCES
C4a-hydroperoxyflavin
FAD
S-oxygenation
allicin
allyl mercaptan
class B monooxygenases
flavin
flavin-dependent monooxygenases
garlic (Allium sativum)
hydroxylation
natural product
natural product biosynthesis
oxidation-reduction (redox)
plant biochemistry
structure-function
sulfenic acid

Structure and function of a flavin-dependent S-monooxygenase from garlic (Allium sativum)

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