THE ACTIVATION GATE OF CYCLIC NUCLEOTIDE-GATED CHANNELS

Project: Research project

Description

DESCRIPTION (provided by applicant): Cyclic nucleotide-gated (CNG) channels play a fundamental role in photoreceptor neurons where they mediate the final step of the signal transduction cascade triggered by absorption of photons. They are Ca2+ permeable, non-selective cation channels that are opened by binding of cyclic nucleotides. Mutations in CNG channel genes are responsible for numerous diseases in the visual system including retinitis pigmentosa (retinal degeneration) and achromatopsia (total color blindness). CNG channels are members of a large family of P-loop containing ion channels that shares considerable common structural and functional similarities. The long-term goal of the proposed experiments is to understand how conformational rearrangements in the channel protein lead to the opening of the CNG channel pore. While recent crystal structures of bacteria P-loop containing channels revealed both the closed and the open pore conformations, many fundamental questions regarding the molecular mechanism of channel gating remain unanswered. What pore structure constitutes the activation gate(s)? What are the structural movements that initiate ion permeation? To address these and related questions in CNG channels, the grant will take particular advantage of our recently developed fluorescence technique (termed Patch-Clamp Fluorometry, or PCF) that is based on site-specific fluorescence labeling of channels in isolated membrane patches. PCF will be used to simultaneously record fluorescence and current signals from the same population of channels in real time, and to directly correlate structural changes in the channel protein to their gating effects. PCF will be combined with fluorescence quenching and fluorescence resonance energy transfer (FRET) to investigate the channel pore structure as well as rearrangements in the structure during CNG channel activation. Findings from these experiments will advance our understanding of the molecular basis for CNG channel gating under physiological as well as disease states.
StatusFinished
Effective start/end date2/1/071/31/10

Funding

  • National Institutes of Health: $184,594.00
  • National Institutes of Health: $218,377.00

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Cyclic Nucleotide-Gated Cation Channels
Fluorescence
Color Vision Defects
Fluorometry
Retinal Degeneration
Fluorescence Resonance Energy Transfer
Retinitis Pigmentosa
Organized Financing
Cyclic Nucleotides
Patch-Clamp Techniques
Ion Channels
Photons
Cations
Signal Transduction
Proteins
Ions
Bacteria
Neurons
Mutation
Membranes

ASJC

  • Medicine(all)