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Article
Identification, Recombinant Expression, and Biochemical Analysis of Putative Secondary Product Glucosyltransferases from Citrus paradisi.
Journal of Agricultural and Food Chemistry (2016)
  • Shivakumar P. Devaiah
  • Daniel K. Owens, East Tennessee State University
  • Mebrahtu B. Sibhatu
  • Tapasree R Sarkar
  • Christy L Strong
  • Venkata K. P. S. Mallampalli
  • Jesephat Asiago
  • Jennifer Cooke
  • Starla Kiser
  • Zhangfan Lin
  • Anye Wamucho
  • Deborah Hayford
  • Bruce E. Williams
  • Peri Loftis
  • Mark Berhow
  • Lee M. Pike
  • Cecilia A. McIntosh, East Tennessee State University
Abstract
Flavonoid and limonoid glycosides influence taste properties as well as marketability of Citrus fruit and products, particularly grapefruit. In this work, nine grapefruit putative natural product glucosyltransferases (PGTs) were resolved by either using degenerate primers against the semiconserved PSPG box motif, SMART-RACE RT-PCR, and primer walking to full-length coding regions; screening a directionally cloned young grapefruit leaf EST library; designing primers against sequences from other Citrus species; or identifying PGTs from Citrus contigs in the harvEST database. The PGT proteins associated with the identified full-length coding regions were recombinantly expressed in Escherichia coli and/or Pichia pastoris and then tested for activity with a suite of substrates including flavonoid, simple phenolic, coumarin, and/or limonoid compounds. A number of these compounds were eliminated from the predicted and/or potential substrate pool for the identified PGTs. Enzyme activity was detected in some instances with quercetin and catechol glucosyltransferase activities having been identified.

Keywords
  • Citrus paradisi,
  • flavonoid,
  • flavor,
  • glucosyltransferase,
  • heterologous expression
Disciplines
Publication Date
March 9, 2016
DOI
10.1021/acs.jafc.5b05430
Citation Information
Devaiah, S. P., Owens, D. K., Sibhatu, M. B., Sarkar, T. R., Strong, C. L., Mallampalli, V. K. P. S., … McIntosh, C. A. (2016). Identification, Recombinant Expression, and Biochemical Analysis of Putative Secondary Product Glucosyltransferases from Citrus paradisi. Journal of Agricultural and Food Chemistry, 64(9), 1957–1969. https://doi.org/10.1021/acs.jafc.5b05430