Proteomics in brain research: potentials and limitations

Abstract

The advent of proteomics techniques has been enthusiastically accepted in most areas of biology and medicine. In neuroscience, a host of applications was proposed ranging from neurotoxicology, neurometabolism, determination of the proteome of the individual brain areas in health and disease, to name a few. Only recently, the limitations of the method have been shown, hampering the rapid spreading of the technology, which in principle consists of two-dimensional gel electrophoresis with in-gel protein digestion of protein spots and identification by mass-spectrometrical approaches or microsequencing. The identification, including quantification using specific software, of brain protein classes, like enzymes, cytoskeleton proteins, heat shock proteins/chaperones, proteins of the transcription and translation machinery, synaptosomal proteins, antioxidant proteins, is a clear domain of proteomics. Furthermore, the concomitant detection of several hundred proteins on a gel allows the demonstration of an expressional pattern, rather generated by a reliable, protein-chemical method than by immunoreactivity, proposed by protein-arrays. An additional advantage is that hitherto unknown proteins, so far only proposed from their nucleic acid structure, designated as hypothetical proteins, can be identified as brain proteins. As to shortcomings and disadvantages of the method we would point to the major problem, the failure to separate hydrophobic proteins. There is so far no way to analyse the vast majority of these proteins in gels. Several other analytical problems need to be overcome, but once the latter problem can be solved, there is nothing to stop the method for a large scale analysis of membrane proteins in neuroscience.

Introduction

Proteomics is the science and methodology of the study of the proteome, i.e. all proteins expressed in a cell or tissue, rather than proteins one by one. The advent of proteomics was a major step forward, comparable to the introduction of molecular biological methods in the past. It fits within the concept that the determination of protein rather than RNA levels has major advantages as it is the proteins that carry out functions. There is a long and unpredictable way from RNA to protein, a fact known to all scientists working with gene hunting methods, like differential display and subtractive hybridization. Huge subtractive libraries have been generated, but when the differentially expressed mRNAs were studied at the protein level, only a small percentage of aberrant transcriptomes could be verified. This does not mean, of course, that proteomics is fully replacing these techniques, but proteomics is a very valuable tool for protein hunting, i.e. comparing cell and tissue proteomes under physiological and pathophysiological conditions. The trend, however, seems to be that once the protein is determined, the transcriptional level is examined to find the underlying mechanism for the increase or decrease of a certain gene product. Moreover, information about the presence of isoforms and post-translational modifications can be obtained by proteomics.

In the following, we give only an outline, an example of a proteomic method that has been used in many studies, and we are aware that much valuable work is not mentioned and many important studies are not cited. The review is written to enable the neuroscientist to catch the spirit of proteomics. After introducing the methodology, we present typical human brain protein maps, identify the protein classes, recommend applications, discuss the shortcomings and limitations of proteomics, and, finally, draw a conclusion.