Synchrotron radiation circular dichroism spectroscopy applied to metmyoglobin and a 4-alpha-helix bundle carboprotein |
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Authors: | Thulstrup Peter W Brask Jesper Jensen Knud J Larsen Erik |
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Affiliation: | Department of Natural Sciences, Royal Veterinary and Agricultural University, Thorvaldsensvej 40, DK-1871 Frederiksberg C, Denmark. pwt@kvl.dk |
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Abstract: | The novel technique, synchrotron radiation-based circular dichroism (SR-CD), has been applied to the study of metmyoglobin and a carboprotein (carbohydrate-based peptide with protein tertiary structure) with 4-alpha-helix bundle structure, as well as a carbopeptide (carbohydrate-based peptide) with a truncated peptide sequence. The use of synchroton radiation (SR) enabled circular dichroism (CD) measurements in the vacuum ultraviolet (VUV) down to 168 nm in D(2)O and 160 nm in 2,2,2-trifluoroethanol (TFE). The band shape in the CD spectra in the low wavelength region was studied, comparing samples with two types of alpha-helical tertiary structure, namely the globin fold and the 4-alpha-helix bundle motif. No significant differences were found between the CD spectra of the alpha-helical samples (metmyoglobin and carboprotein) in D(2)O solution. The use of 2,2,2-TFE (TFE) as solvent clearly alters the VUV CD but the two samples have very similar CD spectra. The solvent-induced denaturing of metmyoglobin in TFE was observed using absorption and CD spectroscopy of the Soret band, with results indicating heme release. The VUV spectrum of TFE-denatured metmyoglobin exhibits dramatic differences in comparison with previous studies of the native enzyme in aqueous solution. The implications of this observation are discussed. |
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Keywords: | SR‐CD synchrotron radiation circular dichroism VUV metmyoglobin carboprotein carbopeptide 2,2,2‐TFE TFE α‐helix protein folding |
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