Functional characterization of the pentapeptide QYNAD on rNav1.2 channels and its NMR structure.
The endogenous pentapeptide QYNAD (Gln-Tyr-Asn-Ala-Asp) is present in human cerebrospinal fluid (CSF), and its concentration is increased in demyelinating diseases. QYNAD was synthesized and its action on the rNav1.2 voltage-gated sodium channel alpha-subunit was studied using whole-cell recordings in a heterologous expression system. The effects were seen only upon equilibration of the peptide in the external bath solution for at least 10 min before the commencement of whole-cell experiments. The steady-state activation curve showed a rightward shift of 10 mV, while the steady-state inactivation curve showed a leftward shift of 5 mV. Frequency-dependent inhibition of the sodium current amplitude was observed at 2-10 Hz, in the presence of external QYNAD, but was not seen when applied internally. Fits of the whole-cell sodium current traces by Hodgkin-Huxley equations revealed subtle changes in the voltage-dependent rate constants governing the transition of the activation and the inactivation gates. Two dimensional NMR spectroscopy revealed the absence of medium and long-range Nuclear Overhauser effects (NOEs), which indicates that the peptide does not adopt any canonical secondary structure in solution. In summary, our studies show that although the pentapeptide QYNAD does not have a defined structure in solution, it has defined actions on the rNav1.2 voltage-gated sodium channel isoform.
Padmashri
R
R
Membrane and Cellular Biophysics, Molecular Biophysics Unit, Indian Institute of Science, 560012 Bangalore, India.
Chakrabarti
K S
KS
Sahal
D
D
Mahalakshmi
R
R
Sarma
S P
SP
Sikdar
S K
SK
D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D009682:Magnetic Resonance Spectroscopy / Q000379:methods; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology*; D062551:NAV1.2 Voltage-Gated Sodium Channel; D009419:Nerve Tissue Proteins / Q000037:antagonists & inhibitors* / Q000502:physiology; D009842:Oligopeptides / Q000737:chemistry* / Q000502:physiology*; D051381:Rats; D015222:Sodium Channels / Q000502:physiology
D062551:NAV1.2 Voltage-Gated Sodium Channel; D009419:Nerve Tissue Proteins; D009842:Oligopeptides; C568251:SCN2A protein, human; C568253:Scn2A protein, rat; D015222:Sodium Channels; C450845:pentapeptide QYNAD