Forskning ved Københavns Universitet - Københavns Universitet

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A novel analytical method for in vivo phosphate tracking

Publikation: Bidrag til tidsskriftTidsskriftartikelForskningfagfællebedømt

  • Hong Gu
  • Sylvie Lalonde
  • Sakiko Okumoto
  • Loren L. Looger
  • Anne Marie Scharff-Poulsen
  • Arthur R. Grossman
  • Jens Kossmann
  • Iver Jakobsen
  • Wolf B. Frommer

Genetically-encoded fluorescence resonance energy transfer (FRET) sensors for phosphate (Pi) (FLIPPi) were engineered by fusing a predicted Synechococcus phosphate-binding protein (PiBP) to eCFP and Venus. Purified fluorescent indicator protein for inorganic phosphate (FLIPPi), in which the fluorophores are attached to the same PiBP lobe, shows Pi-dependent increases in FRET efficiency. FLIPPi affinity mutants cover Pi changes over eight orders of magnitude. COS-7 cells co-expressing a low-affinity FLIPPi and a Na+/Pi co-transporter exhibited FRET changes when perfused with 100µM Pi, demonstrating concentrative Pi uptake by PiT2. FLIPPi sensors are suitable for real-time monitoring of Pi metabolism in living cells, providing a new tool for fluxomics, analysis of pathophysiology or changes of Pi during cell migration.

OriginalsprogEngelsk
TidsskriftFEBS Letters
Vol/bind580
Udgave nummer25
Sider (fra-til)5885-5893
Antal sider9
ISSN0014-5793
DOI
StatusUdgivet - 2006

ID: 8042551