Characterization of recombinant plant cinnamate 4-hydroxylase produced in yeast.

Item request has been placed! ×
Item request cannot be made. ×
loading   Processing Request
  • Additional Information
    • Subject Terms:
    • Abstract:
      Helianthus tuberosus cinnamate 4-hydroxylase (CYP73 or CA4H), a member of the P450 superfamily which catalyses the first oxidative step of the phenylpropanoid pathway in higher plants by transforming cinnamate into p-coumarate, was expressed in the yeast Saccharomyces cerevisiae. The PCR-amplified CA4H open reading frame was inserted into pYeDP60 under the transcriptional control of a galactose-inducible artificial promoter: Engineered S. cerevisiæ strains producing human P450 reductase or normal or overproduced amounts of yeast P450 reductase were transformed to express recombinant CA4H. When grown on galactose, yeast cells produced CA4H holoprotein bound to the endoplasmic reticulum membrane as judged from the reduced iron/carbon monoxide difference spectrum centered at 452 nm and from typical cinnamate 4-hydroxylase activity upon coupling with the different P450 reductases and NADPH. Some CA4H protein was found also addressed to the yeast mitochondria but as a low-activity form. The spectral and kinetic characterizations of the yeast-produced CA4H in different redox protein environments are presented using both assays on yeast microsomal fractions and bioconversions on living ceils. Results indicate that the microsomal system constituted; by the overexpressed yeast P450 reductase and CA4H is characterized by a 1:1 coupling between NADPH oxidation and cinnamate hydroxylation and by one of the highest turnover numbers reported for an NADPH-dependent P450 reaction. Based on spectral perturbation and inhibition studies, coumarate appeared to have no detectable affinity for the enzyme. A possible geometry of the substrate recognition pocket is discussed in the light of these data. [ABSTRACT FROM AUTHOR]
    • Abstract:
      Copyright of European Journal of Biochemistry is the property of Wiley-Blackwell and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)
    • ISSN:
      0014-2956
    • Accession Number:
      10.1111/j.1432-1033.1994.tb18931.x
    • Accession Number:
      12666937
  • Citations
    • ABNT:
      URBAN, P. et al. Characterization of recombinant plant cinnamate 4-hydroxylase produced in yeast. European Journal of Biochemistry, [s. l.], v. 222, n. 3, p. 843–850, 1994. DOI 10.1111/j.1432-1033.1994.tb18931.x. Disponível em: http://search.ebscohost.com/login.aspx?direct=true&site=eds-live&db=hch&AN=12666937. Acesso em: 2 dez. 2020.
    • AMA:
      Urban P, Werck-Reichhart D, Teutsch HG, et al. Characterization of recombinant plant cinnamate 4-hydroxylase produced in yeast. European Journal of Biochemistry. 1994;222(3):843-850. doi:10.1111/j.1432-1033.1994.tb18931.x
    • APA:
      Urban, P., Werck-Reichhart, D., Teutsch, H. G., Durst, F., Regnier, S., Kazmaier, M., & Pompon, D. (1994). Characterization of recombinant plant cinnamate 4-hydroxylase produced in yeast. European Journal of Biochemistry, 222(3), 843–850. https://doi.org/10.1111/j.1432-1033.1994.tb18931.x
    • Chicago/Turabian: Author-Date:
      Urban, Philippe, Danièle Werck-Reichhart, Hermann G. Teutsch, Francis Durst, Sylvie Regnier, Michael Kazmaier, and Denis Pompon. 1994. “Characterization of Recombinant Plant Cinnamate 4-Hydroxylase Produced in Yeast.” European Journal of Biochemistry 222 (3): 843–50. doi:10.1111/j.1432-1033.1994.tb18931.x.
    • Harvard:
      Urban, P. et al. (1994) ‘Characterization of recombinant plant cinnamate 4-hydroxylase produced in yeast’, European Journal of Biochemistry, 222(3), pp. 843–850. doi: 10.1111/j.1432-1033.1994.tb18931.x.
    • Harvard: Australian:
      Urban, P, Werck-Reichhart, D, Teutsch, HG, Durst, F, Regnier, S, Kazmaier, M & Pompon, D 1994, ‘Characterization of recombinant plant cinnamate 4-hydroxylase produced in yeast’, European Journal of Biochemistry, vol. 222, no. 3, pp. 843–850, viewed 2 December 2020, .
    • MLA:
      Urban, Philippe, et al. “Characterization of Recombinant Plant Cinnamate 4-Hydroxylase Produced in Yeast.” European Journal of Biochemistry, vol. 222, no. 3, June 1994, pp. 843–850. EBSCOhost, doi:10.1111/j.1432-1033.1994.tb18931.x.
    • Chicago/Turabian: Humanities:
      Urban, Philippe, Danièle Werck-Reichhart, Hermann G. Teutsch, Francis Durst, Sylvie Regnier, Michael Kazmaier, and Denis Pompon. “Characterization of Recombinant Plant Cinnamate 4-Hydroxylase Produced in Yeast.” European Journal of Biochemistry 222, no. 3 (June 15, 1994): 843–50. doi:10.1111/j.1432-1033.1994.tb18931.x.
    • Vancouver/ICMJE:
      Urban P, Werck-Reichhart D, Teutsch HG, Durst F, Regnier S, Kazmaier M, et al. Characterization of recombinant plant cinnamate 4-hydroxylase produced in yeast. European Journal of Biochemistry [Internet]. 1994 Jun 15 [cited 2020 Dec 2];222(3):843–50. Available from: http://search.ebscohost.com/login.aspx?direct=true&site=eds-live&db=hch&AN=12666937