Refining protein amide I spectrum simulations with simple yet effective electrostatic models for local wavenumbers and dipole derivative magnitudes.

Analysis of the amide I band of proteins is probably the most wide-spread application of bioanalytical infrared spectroscopy. Although highly desirable for a more detailed structural interpretation, a quantitative description of this absorption band is still difficult. This work optimized several electrostatic models with the aim to reproduce the effect of the protein environment on the intrinsic wavenumber of a local amide I oscillator. We considered the main secondary structures - α-helices, parallel and antiparallel ß-sheets - with a maximum of 21 amide groups. The models were based on the electric potential and/or the electric field component along the CO bond at up to four atoms in an amide group. They were bench-marked by comparison to Hessian matrices reconstructed from density functional theory calculations at the BPW91, 6-31G** level. The performance of the electrostatic models depended on the charge set used to calculate the electric field and potential. Gromos and DSSP charge sets, used in common force fields, were not optimal for the better performing models. A good compromise between performance and the stability of model parameters was achieved by a model that considered the electric field at the positions of the oxygen, nitrogen, and hydrogen atoms of the considered amide group. The model describes also some aspects of the local conformation effect and performs similar on its own as in combination with an explicit implementation of the local conformation effect. It is better than a combination of a local hydrogen bonding model with the local conformation effect. Even though the short-range hydrogen bonding model performs worse, it captures important aspects of the local wavenumber sensitivity to the molecular surroundings. We improved also the description of the coupling between local amide I oscillators by developing an electrostatic model for the dependency of the dipole derivative magnitude on the protein environment.

On the Secondary Structure of Silk Fibroin Nanoparticles Obtained Using Ionic Liquids: An Infrared Spectroscopy Study.

Silk fibroin from Bombyx mori caterpillar is an outstanding biocompatible polymer for the production of biomaterials. Its impressive combination of strength, flexibility, and degradability are related to the protein's secondary structure, which may be altered during the manufacture of the biomaterial. The present study looks at the silk fibroin secondary structure during nanoparticle production using ionic liquids and high-power ultrasound using novel infrared spectroscopic approaches. The infrared spectrum of silk fibroin fibers shows that they are composed of 58% ß-sheet, 9% turns, and 33% irregular and/or turn-like structures. When fibroin was dissolved in ionic liquids, its amide I band resembled that of soluble silk and no ß-sheet absorption was detected. Silk fibroin nanoparticles regenerated from the ionic liquid solution exhibited an amide I band that resembled that of the silk fibers but had a reduced ß-sheet content and a corresponding higher content of turns, suggesting an incomplete turn-to-sheet transition during the regeneration process. Both the analysis of the experimental infrared spectrum and spectrum calculations suggest a particular type of ß-sheet structure that was involved in this deficiency, whereas the two other types of ß-sheet structure found in silk fibroin fibers were readily formed.

The Amide I Spectrum of Proteins-Optimization of Transition Dipole Coupling Parameters Using Density Functional Theory Calculations.

The amide I region of the infrared spectrum is related to the protein backbone conformation and can provide important structural information. However, the interpretation of the experimental results is hampered because the theoretical description of the amide I spectrum is still under development. Quantum mechanical calculations, for example, using density functional theory (DFT), can be used to study the amide I spectrum of small systems, but the high computational cost makes them inapplicable to proteins. Other approaches that solve the eigenvalues of the coupled amide I oscillator system are used instead. An important interaction to be considered is transition dipole coupling (TDC). Its calculation depends on the parameters of the transition dipole moment. This work aims to find the optimal parameters for TDC in three major secondary structures: α-helices, antiparallel ß-sheets, and parallel ß-sheets. The parameters were suggested through a comparison between DFT and TDC calculations. The comparison showed a good agreement for the spectral shape and for the wavenumbers of the normal modes for all secondary structures. The matching between the two methods improved when hydrogen bonding to the amide oxygen was considered. Optimal parameters for individual secondary structures were also suggested.

Insight into the internal structure of amyloid-ß oligomers by isotope-edited Fourier transform infrared spectroscopy.

The internal structure of amyloid-ß (Aß) oligomers was investigated with isotope-edited Fourier transform infrared spectroscopy. Homo-oligomers of Aß40 and Aß42 were prepared from unlabeled and 13C, 15N-labeled monomeric Aß and from mixtures of these. For the unlabeled peptides, two main bands were observed in 2H2O at 1685 and 1622 cm-1 for Aß40 and at 1685 and 1626 cm-1 for Aß42. These band positions indicate that the number of strands per sheet is at least four. The obtained experimental amide I spectra were simulated using a number of structural models (antiparallel ß-sheets, ß-barrels and a dodecamer structure). According to experiments and calculations, the main 13C-band shifts down at increasing molar ratio of labeled peptides. This shift occurs when vibrational coupling becomes possible between 13C-amide groups in close-by strands. It is small, when intervening 12C-strands increase the distance between 13C-strands; it is large, when many neighboring strands are labeled. The shift depends on the internal structure of the peptides within the oligomers, i.e. on the building block that each peptide molecule contributes to the ß-sheets of the oligomers. The shift is largest, when individual peptides contribute just a single strand surrounded by strands from other peptide molecules. It is smaller when each molecule forms two or three adjacent strands. As indicated by a comparison between experiment and computation, the number of adjacent ß-strands per peptide molecule is two for Aß40 oligomers and two or more for Aß42 oligomers. Our results are well explained by regular, antiparallel ß-sheets or ß-barrels.

Amyloid ß-peptides 1-40 and 1-42 form oligomers with mixed ß-sheets.

Two main amyloid-ß peptides of different length (Aß40 and Aß42) are involved in Alzheimer's disease. Their relative abundance is decisive for the severity of the disease and mixed oligomers may contribute to the toxic species. However, little is know about the extent of mixing. To study whether Aß40 and Aß42 co-aggregate, we used Fourier transform infrared spectroscopy in combination with 13C-labeling and spectrum calculation and focused on the amide I vibration, which is sensitive to backbone structure. Mixtures of monomeric labeled Aß40 and unlabeled Aß42 (and vice versa) were co-incubated for ∼20 min and their infrared spectrum recorded. The position of the main 13C-amide I' band shifted to higher wavenumbers with increasing admixture of 12C-peptide due to the presence of 12C-amides in the vicinity of 13C-amides. The results indicate that Aß40 and Aß42 form mixed oligomers with a largely random distribution of Aß40 and Aß42 strands in their ß-sheets. The structures of the mixed oligomers are intermediate between those of the pure oligomers. There is no indication that one of the peptides forces the backbone structure of its oligomers on the other peptide when they are mixed as monomers. We also demonstrate that isotope-edited infrared spectroscopy can distinguish aggregation modulators that integrate into the backbone structure of their interaction partner from those that do not. As an example for the latter case, the pro-inflammatory calcium binding protein S100A9 is shown not to incorporate into the ß-sheets of Aß42.