Nanotechnologies in proteomics

Abstract

Progress in proteomic researches is largely determined by development and implementation of new methods for the revelation and identification of proteins in biological material in a wide concentration range (from 10 23 M to single molecules). The most perspective approaches to address this problem involve (i) nanotechnological physicochemical procedures for the separation of multicomponent protein mixtures; among these of particular interest are biospecific nanotechnological procedures for selection of proteins from multicomponent protein mixtures with their subsequent concentration on solid support; (ii) identification and counting of single molecules by use of molecular detectors. The prototypes of biospecific nanotechnological procedures, based on the capture of ligand biomolecules by biomolecules of immobilized ligate and the concentration of the captured ligands on appropriate surfaces, are well known; these are affinity chromatography, magnetic biobeads technology, different biosensor methods, etc. Here, we review the most promising nanotechnological approaches for selection of proteins and kinetic characterization of their complexes based on these biospecific methods with subsequent MS/MS identification of proteins and protein complexes. Two major groups of methods for the analysis and identification of individual molecules and their complexes by use of molecular detectors will be reviewed: scanning probe microscopy (SPM) (including atomic-force microscopy) and cryomassdetector technology.

AFM fishing nanotechnology is the way to reverse the Avogadro number in proteomics

“…That is why the situation in proteomics is much more complicated than that in genomics, where PCR is widely adopted for the replication of lowabundant molecules. It appears that decisive successes in proteomics will only be gained when the CSL is lowered sufficiently to allow the investigator to detect one molecule in 1 L of biomaterial (10 224 M) and thus approach the reverse Avogadro number (6610 23 molecules?mol 21 ) 21 .…”

“…The required sensitivity (i.e., the ability to register a single molecule in 1 L of biological material) may be achieved by the use of molecular detectors for the analysis of surfaces, which have been enriched by molecules from biological material [21]. The technology involves two stages: biospecific fishing onto the chip surface and the registration of "fished-out" objects by the use of molecular detectors.…”

“…A most promising molecular detector is one based on the atomic force microscope [21]. At the same time, other detecting devices cannot be excluded such as other types of molecular scanning microscopes [22] or electrochemical detectors on nanowires [23] and nanopores [24].…”

“…Analysis of the scanned images makes it possible to calculate the ratio of the number of single molecules to the number of molecular complexes. If antibodies or aptamers to certain proteins are used as immobilized molecules, we may identify their complexes with appropriate antigens and, hence, estimate the total number of antigen molecules in solution [21].…”

“…After the incubation the biochip was washed and the AFM scanning of its surface was carried out. The AFM measurements were made in air using tapping mode on a multimode "NTEGRA" atomic force microscope (NT-MDT, Moscow, Russia) as described in [21]. Cantilevers NSG 10 supplied by NT MDT were used.…”