An extract of bark from the tropical rainforest plant Byrsonima crassifolia was screened for inhibition of diubiquitin formation by the human ubiquitin-conjugating enzyme E2-25K. Activity assays with both the full-length enzyme and a truncated, active catalytic UBC domain revealed that the extract contained inhibitory properties. Separation of the extract into individual components and additional screens identified vitexin as the active inhibitor. An IC50 for vitexin was calculated to be approximately 0.5 mM. Molecular modeling simulations were used to predict the mode of inhibition and NMR spectra were used to confirm the binding site of vitexin to E2-25K.
HaasALRoseIA. (1982) The mechanism of ubiquitin activating enzyme. A kinetic and equilibrium analysis. Journal of Biological Chemistry, 257, 10329–10337.
2.
HershkoA, CiechanoverA. (1998) The ubiquitin system. Annual Review of Biochemistry, 67, 425–479.
3.
CiechanoverA. (1998) The ubiquitin-proteasome pathway: on protein death and cell life. Embo Journal, 17, 7151–7160.
4.
ChenZ, PickartCM. (1990) A 25-kilodalton ubiquitin carrier protein (E2) catalyzes multi-ubiquitin chain synthesis via lysine 48 of nubiquitin. Journal of Biological Chemistry, 265, 21835–21842.
5.
WilsonRCHughesRCFlattJWMeehanEJNgJDTwiggPD. (2009) The crystal structure of full-length ubiquitin-conjugating enzyme E2-25K (huntingtin-interacting protein 2). Acta Crystallographica, F65, 440–444.
6.
WilsonRCEdmondsonSPFlattJWHelmsKMTwiggPD. (2011) The E2-25K Ubiquitin-associated (UBA) Domain Aids in Polyubiquitin Chain Synthesis and Linkage Specificity. Biochemical and Biophysical Research Communications, 405, 662–666.
7.
OuteiroTFTetzlaffJ. (2007) Mechanisms of Disease II: Cellular Protein Quality Control. Seminars in Pediatric Neurology, 14, 15–25.
8.
FliermanD. (2006) E2-25K mediates US11-triggered retro-translocation of MHC class I heavy chains in a permeabilized cell system. Proceedings of the National Academy of Sciences of the United States of America, 103, 11589–11594.
KalchmanMAGrahamRKXiaG, KoideHBHodgsonJGGrahamKCGoldbergYPGietzRDPickartCMHaydenMR. (1996) Huntingtin is ubiquitinated and interacts with a specific ubiquitin-conjugating enzyme. Journal of Biological Chemistry, 271, 19385–19394.
11.
de PrilR, FischerDFRoosRAC, van LeeuwenFW. (2007) Ubiquitin-conjugating enzyme E2-25K increases aggregate formation and cell death in polyglutamine diseases. Molecular and Cellular Neurosciences, 34, 10–19.
12.
SongS, KimSYHongYMJoDGLeeJYShimSMChungCWSeoSJYooYJKohJYLeeMCYatesAJIchijoH, JungYK. (2003) Essential role of E2-25K/Hip-2 in mediating amyloid-beta neurotoxicity. Molecular Cell, 12, 553–563.
13.
SongS, JungYK. (2004) Alzheimer's disease meets the ubiquitin-proteasome system. Trends in Molecular Medicine, 10(11), 565–570.
14.
MortonJF. (1981) Atlas of Medicinal Plants of Middle America, Vol I. Charles C. Thomas Publisher, Springfield, Illinois, pp 412–414.
15.
DukeJABogenschutz-GodwinMJOttesenAR. (2009) Duke's Handbook of Medicinal Plants of Latin America. CRC Press, Boca Raton, Florida, pp 136–138.
16.
MaldiniM, SosaS, MontoroP, GiangasperoP, BalickMJPizzaC, Della LogiaR. (2009) Screening of the topical anti-inflammatory activity of the bark of Acacia cornigera Willdenow, Byrsonima crassifolia Kunth, Sweetia panamensis Yakovlev and the leaves of Sphagneticola trilobata Hitchcock. Journal of Ethnopharmacology, 122, 430–433.
17.
SouzaJNS, SilvaEMLoirA, ReesJFRogezH, LarondelleY. (2008) Antioxidant capacity of four polyphenol-rich Amazonian plant extracts: A correlation study using chemical and biological in vitro assays. Food Chemistry, 106, 331–339.
18.
BejarE, AmarquayeA, CheCTMaloneMHFongHHS. (1995) Constituents of Byrsonima crassifolia and their spasmogenic activity. International Journal of Pharmacognosy, 33, 25–32.
19.
GeissF, HeinrichM, HunklerD, RimplerH. (1995) Proanthocyanidins with (+)-epicatechin units from Byrsonima crassifolia bark. Phytochemistry, 39, 635–643.
20.
SilvaEMSouzaJNS, ReesJFLarondelleY. (2007) Antioxidant activities and polyphenolic contents of fifteen selected plant species from the Amazonian region. Food Chemistry, 101, 1012–1018.
21.
VazquezE, MartinezEMCogordanJADelgadoG. (2002) Triterpenes, phenols, and other constituents from the leaves of Ochroma pyramidale (Balsa wood, Bombacaceae). Preferred conformations of 8-C-β-D-glucopyranosyl-apigenin (vitexin). Revista de la Sociedad Quimica de Mexico, 46, 254–258.
22.
Dictionary of Natural Products. (2010), v. 19:1. CRC Press, Boca Raton, Florida.
RagoneMISellaM, ConfortiP, VolontéMGConsoliniAE. (2007) The spasmolytic effects of Aloysia citriodora, Palau (South American cedrón) is partially due to its vitexin but not isovitexin on rat duodenums. Journal of Ethnopharmacology, 113, 258–266.
25.
GaitanE, CookseyRCLeganJ, LindsayRH. (1995) Antithyroid effects in vivo and in vitro of vitexin: A C-glucosylflavone in millet. Clinical Endrocrinology & Metabolism, 80, 1144–1147.
ChoiHJEunJSKimBGKimSYJeonH, SohY. (2006) Vitexin, an HTF-1α inhibitor, has anti-metastatic potential in PC12 cells. Molecules and Cells, 22, 291–299.
29.
KoS, KangGBSongSMLeeJGShinDYYunJHShengY, CheongC, JeonYHJungYKArrowsmithCHAvvakumovGVDhe-PaganonS, YooYJEomSHLeeW. (2010) Structural basis of E2-25K/UBB+1 interaction leading to proteasome inhibition and neurotoxicity. Journal of Biological Chemistry, 285, 36070–36080.
30.
HuangDTPaydarA, ZhuangM, WaddellMBHoltonJMSchulmanBA. (2005) Structural Basis for Recruitment of Ubc12 by an E2 Binding Domain in NEDD8′s E1. Molecular Cell, 17, 341–350.
31.
ShukerSBHajdukPJMeadowsRPFesikSW. (1996) Discovering high-affinity ligands for proteins: SAR by NMR. Science, 274, 1531–1534.
32.
TsukamotoS, TakeuchiT, RotinsuluH, MangindaanREP, van SoestRWM, UkaiK, KobayashiH, NamikoshiM, OhtaT, YokosawaH. (2008) Leucettamol A: A new inhibitor of Ubc13-Uev1A interaction isolated from a marine sponge, Leucetta aff. microrhaphis. Bioorganic & Medicinal Chemistry Letters, 18, 6319–6320.
33.
KayL, KeiferP, SaarinenT. (1992) Pure absorption gradient enhanced heteronuclear single quantum correlation spectroscopy with improved sensitivity. Journal of the American Chemical Society, 114, 10663–10665.
34.
MarkleyJLBaxA, ArataY, HilbersCWKapteinR, SykesBDWrightPEWuthrichK. (1998) Recommendations for the presentation of NMR structures of proteins and nucleic acids. IUPAC-IUBMB-IUPAB Inter-Union Task Group on the Standardization of Data Bases of Protein and Nucleic Acid Structures Determined by NMR Spectroscopy. Journal of Biomolecular NMR, 12, 1– 23.
35.
DelaglioF, GrzesiekS, VuisterGWZhuG, PfeiferJ, BaxA. (1995) NMRPipe: a multidimensional spectral processing system based on UNIX pipes. Journal of Biomolecular NMR, 6, 277–293.
36.
JohnsonBA. (2004) Using NMRView to visualize and analyze the NMR spectra of macromolecules. Methods in Molecular Biology, 278, 313–352.
37.
PiotrowskiJ, BealR, HoffmanL, WilkinsonKDCohenREPickartCM. (1997) Inhibition of the 26S proteasome by polyubiquitin chains synthesized to have defined lengths. Journal of Biological Chemistry, 272, 23712–23721.
38.
Molegro Virtual Docker v. 4.3 (2011) Molegro ApS. Aarhus, Denmark.
39.
ThomsenR, ChristensenMH. (2006) MolDock: A new technique for high-accuracy molecular docking. Journal of Medicinal Chemistry, 49, 3315–3321.
