Microbial metalloproteomes are largely uncharacterized.

Metal ion cofactors afford proteins virtually unlimited catalytic potential, enable electron transfer reactions and have a great impact on protein stability. Consequently, metalloproteins have key roles in most biological processes, including respiration (iron and copper), photosynthesis (manganese) and drug metabolism (iron). Yet, predicting from genome sequence the numbers and types of metal an organism assimilates from its environment or uses in its metalloproteome is currently impossible because metal coordination sites are diverse and poorly recognized.

Novel multiprotein complexes identified in the hyperthermophilic archaeon Pyrococcus furiosus by non-denaturing fractionation of the native proteome.

Virtually all cellular processes are carried out by dynamic molecular assemblies or multiprotein complexes, the compositions of which are largely undefined. They cannot be predicted solely from bioinformatics analyses nor are there well defined techniques currently available to unequivocally identify protein complexes (PCs). To address this issue, we attempted to directly determine the identity of PCs from native microbial biomass using Pyrococcus furiosus, a hyperthermophilic archaeon that grows optimally at 100 degrees C, as the model organism.

A general model-based approach to two-dimensional infrared correlation spectroscopy incorporating the global phase angle.

We describe a theoretical framework for a model-based approach to two-dimensional correlation spectroscopy that is generally applicable to any arbitrary model function. The method is based on the correlation between spectral data and a set of model waveforms with a varying correlation index, the global phase angle Theta. When experimental spectral intensity variations are expressed as sinusoidal, exponential, Lorentzian, or quadratic functions, the proposed approach allows us to estimate the quantitative values of the target parameters in those expressions.

Rapid and sensitive detection of respiratory virus molecular signatures using a silver nanorod array SERS substrate.

A spectroscopic assay based on surface enhanced Raman scattering (SERS) using silver nanorod array substrates has been developed that allows for rapid detection of trace levels of viruses with a high degree of sensitivity and specificity. This novel SERS assay can detect spectral differences between viruses, viral strains, and viruses with gene deletions in biological media. The method provides rapid diagnostics for detection and characterization of viruses generating reproducible spectra without viral manipulation.

Polarized surface enhanced Raman and absorbance spectra of aligned silver nanorod arrays.

Polarized surface-enhanced Raman scattering (SERS) and UV-vis absorbance spectra were measured for a nonplanar Ag nanorod array substrate prepared by oblique angle vapor deposition. The anisotropy of the SERS polarization was shown to differ from that of the polarized UV-vis absorbance. The maximum SERS intensity was observed in the polarization direction perpendicular to the long axis of the Ag nanorods, while the UV-vis absorbance was strongly polarized along the direction of the long axis of the nanorod array.

Structure and properties of phospholipid-peptide monolayers containing monomeric SP-B(1-25) II. Peptide conformation by infrared spectroscopy.

The conformation and orientation of synthetic monomeric human sequence SP-B(1-25) (mSP-B(1-25)) was studied in films with phospholipids at the air-water (A/W) interface by polarization modulation infrared reflectance absorption spectroscopy (PM-IRRAS). Modified two-dimensional infrared (2D IR) correlation analysis was applied to PM-IRRAS spectra to define changes in the secondary structure and rates of reorientation of mSP-B(1-25) in the monolayer during compression. PM-IRRAS spectra and 2D IR correlation analysis showed that, in pure films, mSP-B(1-25) had a major alpha-helical conformation plus regions of beta-sheet structure.

Structure and properties of phospholipid-peptide monolayers containing monomeric SP-B(1-25) I. Phases and morphology by epifluorescence microscopy.

Epifluorescence microscopy was used to study the structure and phase behavior of phospholipid films containing a human-sequence monomeric SP-B(1-25) synthetic peptide (mSP-B(1-25)). Measurements were done directly at the air-water (A/W) interface on films in a Langmuir-Whilhelmy balance coupled to a fluorescence microscope and real-time detection system to yield an approximate optical resolution of 1 mum. Fluorescence was achieved by laser excitation of 2-(4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-diaza-s-indacene-3-dodecanoyl)-1-hexadecanoyl-sn-glycero-3-PC (BODIPY-PC, concentration View Article and Find Full Text PDF

Deacylated pulmonary surfactant protein SP-C transforms from alpha-helical to amyloid fibril structure via a pH-dependent mechanism: an infrared structural investigation.

Bovine pulmonary surfactant protein C (SP-C) is a hydrophobic, alpha-helical membrane-associated lipoprotein in which cysteines C4 and C5 are acylated with palmitoyl chains. Recently, it has been found that the alpha-helix form of SP-C is metastable, and under certain circumstances may transform from an alpha-helix to a beta-strand conformation that resembles amyloid fibrils. This transformation is accelerated when the protein is in its deacylated form (dSP-C).

Effect of hydrophobic surfactant proteins SP-B and SP-C on phospholipid monolayers. Protein structure studied using 2D IR and beta correlation analysis.

We have applied two-dimensional infrared (2D IR) and betanu correlation spectroscopy to in-situ IR spectroscopy of pulmonary surfactant proteins SP-B and SP-C in lipid-protein monolayers at the air-water interface. For both SP-B and SP-C, a statistical windowed autocorrelation method identified two separate surface pressure regions that contained maximum amide I intensity changes: 4-25 mN/m and 25-40 mN/m. For SP-C, 2D IR and betanu correlation analyses of these regions indicated that SP-C adopts a variety of secondary structure conformations, including alpha-helix, beta-sheet, and an intermolecular aggregation of extended beta-sheet structure.