Synthesis and secondary structure of loop 4 of myelin proteolipid protein: effect of a point mutation found in Pelizaeus-Merzbacher disease.
To study the effects of a point mutation found in Pelizaeus-Merzbacher disease (PMD) on the physicochemical and structural properties of the extracellular loop 4 of the myelin proteolipid protein (PLP), we synthesized the peptide PLP(181-230)Pro215 and one mutant PLP(181-230)Ser215 with regioselective formation of the two disulphide bridges Cys200-Cys219 and Cys183-Cys227. As conventional amino acid building blocks failed to give crude peptides of good quality we had to optimize the synthesis by introducing pseudoproline dipeptide building blocks during the peptide elongation. In peptide Pro215 the first bridge Cys200-Cys219 was obtained after air oxidation, but in peptide Ser215 because of aggregation, dimethyl sulfoxide (DMSO) oxidation had to be used. The second bridge Cys183-Cys227 was obtained by iodine oxidation of both Cys (acetamidomethyl, Acm)-protected peptides. The secondary structures of the parent and mutant loops were analysed by circular dichroism (CD) in the presence of trifluoroethanol (TFE) and sodium dodecyl sulphate (SDS) as a membrane mimetic. Analysis of the spectra showed that the content of alpha-helix and beta-sheet varied differently for both peptides in TFE and SDS solutions, demonstrating the sensitivity of their conformation to the environment and the differences in their secondary structure. The ability of both peptides to insert into the SDS micelles was assayed by intrinsic tryptophan fluorescence.
Trifilieff
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Laboratoire de Chimie Organique des Substances Naturelles, UMR 7123 CNRS/Université Louis Pasteur Strasbourg, Centre de Neurochimie, 5, rue Blaise Pascal, 67084 Strasbourg Cedex, France. trif@chimie.u-strasbg.fr
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D002942:Circular Dichroism; D004121:Dimethyl Sulfoxide; D006801:Humans; D007455:Iodine / Q000378:metabolism; D008823:Micelles; D008969:Molecular Sequence Data; D018991:Myelin Proteolipid Protein / Q000138:chemical synthesis* / Q000737:chemistry; D010084:Oxidation-Reduction; D020371:Pelizaeus-Merzbacher Disease / Q000235:genetics*; D010446:Peptide Fragments; D010455:Peptides / Q000138:chemical synthesis / Q000737:chemistry; D017354:Point Mutation; D011392:Proline / Q000737:chemistry; D017433:Protein Structure, Secondary; D012694:Serine / Q000737:chemistry; D012967:Sodium Dodecyl Sulfate / Q000737:chemistry; D014270:Trifluoroethanol / Q000737:chemistry
D008823:Micelles; D018991:Myelin Proteolipid Protein; D010446:Peptide Fragments; D010455:Peptides; D012967:Sodium Dodecyl Sulfate; D012694:Serine; D014270:Trifluoroethanol; D007455:Iodine; D011392:Proline; D004121:Dimethyl Sulfoxide