Application of surface enhanced Raman spectroscopy to investigate Permeability of connexin 43 Hemichannels in lipid bilayers And DNA mutations

Abstract

Hemichannels formed by Connexin 43 Cx43 are overexpressed in newlineSf9 insect cells and reconstituted in planar bilayers and large unilamellar newlineVesicles We characterized the permeability of these reconstituted newlinehemichannels to different fluorescent tracers and non fluorescent cytoplasmic newlinemolecules using patch clamp fluorescence and Surface Enhanced Raman newlineSpectroscopy techniques Patch clamp recording of the hemichannel activity newlineafter reconstitution in planar phospholipid bilayers combined with surface newlineenhanced Raman spectroscopy SERS measurements is used to quantify the newlineratio of the permeability of Rhodamine 6G R6G molecules with respect to newlineK ions Hemichannel permeability of a variety of fluorescent probes ranging newlinefrom R6G to short single stranded oligonucleotides is also estimated in giant newlineunilamellar vesicles GUV from fluorescence and SERS measurements newlineHemichannel permeability of a non fluorescent molecule ATP is measured newlinefor the first time employing SERS technique and results are reported Attempt newlineis made to correlate the structural changes accompanying mutations of DNA newlinefragments coding for Cx26 protein with SERS spectra Results of these newlineexperiments could be of relevance for pre screening the DNA of individuals newlinein families genetically pre disposed to certain Connexin related diseases newline