NMR studies of the protonation states of pyridoxal-5′-phosphate in water

Monique Chan-Huot, Christiane Niether, Shasad Sharif, Peter M. Tolstoy, Michael D. Toney, Hans Heinrich Limbach

Research output: Contribution to journalArticle

20 Scopus citations

Abstract

We have measured the 13C NMR spectra of the cofactor pyridoxal-5′-phosphate (vitamin B6, PLP) at 278 K in aqueous solution as a function of pH. By 13C enrichment of PLP in the C-4′ and C-5′ positions we were able to measure spectra down to pH 1. From the dependence of the 13C chemical shifts on pH, the pKa values of PLP could be determined. In particular, the heretofore uncharacterized protonation state of PLP, in which the phosphate group as well as the pyridine ring and the phenolic groups are fully protonated, has been analyzed. The corresponding pKa value of 2.4 indicates that the phosphate group is solely involved in the first deprotonation step. The 15N chemical shifts of the pyridine ring of PLP published previously are in good agreement with the new results. These shifts contain information about the tautomerism of the different protonation states of PLP. The implications of these findings for the biological function of PLP are discussed.

Original languageEnglish (US)
Pages (from-to)282-289
Number of pages8
JournalJournal of Molecular Structure
Volume976
Issue number1-3
DOIs
StatePublished - Jul 15 2010

Keywords

  • C Labeling
  • C NMR
  • N NMR
  • Protonation states
  • Pyridoxal-5′-phosphate
  • Vitamin B

ASJC Scopus subject areas

  • Spectroscopy
  • Analytical Chemistry
  • Inorganic Chemistry
  • Organic Chemistry

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    Chan-Huot, M., Niether, C., Sharif, S., Tolstoy, P. M., Toney, M. D., & Limbach, H. H. (2010). NMR studies of the protonation states of pyridoxal-5′-phosphate in water. Journal of Molecular Structure, 976(1-3), 282-289. https://doi.org/10.1016/j.molstruc.2010.03.032