ABSTRACT
Our goal was to compare two popular analytical techniques used nowadays in proteomic investigations for proteins/peptides sequencing and identification, a widely used nanoLC-MS/MS approach applied in the bottom-up proteomics and electron transfer dissociation/proton transfer reaction fragmentation preferably used when top-down strategy is applied. Comparison was carried out with the aid of the ESI-quadrupole ion-trap instrument using the following criteria: total time of analysis including sample preparation, sequence coverage, Mascot scoring, capability to detect modifications, quality of the results as a function of protein molecular weight and sample consumption.
Subject(s)
Chromatography, Liquid/methods , Mass Spectrometry/methods , Proteins/chemistry , Proteomics/instrumentation , Proteomics/methods , Sequence Analysis, Protein/methods , Amino Acid Sequence , Animals , Methanol/chemistry , Molecular Sequence Data , Molecular Weight , Peptides/analysis , Peptides/chemistry , Proteins/analysis , Tandem Mass SpectrometryABSTRACT
Gel electrophoresis serves as a basic analytical tool in the proteomic studies. However, processing of gel electrophoretic images is still the main bottleneck of data analysis, and there is an increasing need for the fully automated approaches. The proposed start-to-end strategy of analyzing the gel images consists of chemometric tools, which allow their effective preprocessing, automatic warping, and data modeling. The image preprocessing techniques: denoising in the wavelet domain and the penalized asymmetric least squares approach for the background estimation are proposed. Matching of images is based on fuzzy warping of features, extracted from the gel images. For the classification or calibration purpose, multivariate approaches such, as partial least squares (PLS) or kernel-PLS methods are used. Performance of the proposed strategy is demonstrated on the real set of the two-dimensional gel images.
Subject(s)
Electrophoresis, Gel, Two-Dimensional/methods , Animals , Cells, Cultured , Models, Theoretical , RatsABSTRACT
Proteome is a natural consequence of the post-genome era when the HUGO project (Human Genome Organization) has almost been completed. Here, a specifically aimed proteome in drug dependence--morphinome, is described, including tasks, strategies and pitfalls of the methodology.
