Structure characterization of recombinant deacylated human pulmonary surfactant protein C and its mutants using infrared spectroscopy and 2D correlation analysis
MetadataShow full item record
Various FTIR spectroscopy sampling techniques are employed to characterize the structures of recombinant deacylated human pulmonary surfactant protein C (rdhSP-C) and its recombinant mutants in organic solvent, or in phospholipid environment. Two-dimensional (2D) correlation analysis techniques are applied to provide improved resolution enhancement for structure determination. Chapter 3 to Chapter 7 are based on individual manuscripts to be submitted. Chapter 3 and Chapter 4 describe the investigations of a pH-dependent mechanism for misfolding of wild type rdhSP-C and its mutants using FTIR transmission spectroscopy and 2D correlation analysis. In Chapter 3, various 2D correlation analysis techniques are explored. Compared to generalized 2D correlation analysis, model-based 2D correlation analysis proves to be a straightforward and quantitative tool in protein structure determination. A pronounced ± to ² structure conversion is manifested for wild type rdhSP-C. In Chapter 4, as site-directed mutageneses moving from the N-terminus to the poly-valine segment, rdhS-C mutants unfold into ²-structures and unordered structures respectively. Chapter 5 describes the surface pressure induced structure changes of wild type rdhSP-C and one of the mutants (K11Q/R12Q) in phospholipid monolayers at the air/water interface. The combination of PM-IRRAS with model-based 2D correlation analysis proves to be very effective and suitable for investigation of model biomembrane system at the air/water interface. Chapter 6 describes the investigation of thermal induced structure changes of rdhSP-C Mut4 (K11Q/R12Q) in planar supported phospholipid bilayers. In addition, polarization ATR-FTIR is applied to elucidate the orientations of the phospholipid acyl chains and the ²-sheet conformation of Mut4 respectively. Systematic studies on rdhSP-C and its mutants indicate that different structure motifs of rdhSP-C may be involved in the unfolding and misfolding process. In addition, environmental changes, i.e. from monolayer to bilayers, may also contribute to the formation of ²-aggregations of rdhSP-C. Chapter 7 describes the methodology using silver nanorods (AgNRs) based intrinsic surface enhanced Raman spectroscopy (SERS) and chemometrics in the differentiation of rotavirus strains.