The microbial proteome database — an automated laboratory catalogue for monitoring protein expression in bacteria

Abstract

Laboratories devoted to high-throughput characterisation of purified proteins arrayed via two-dimensional (2-D) gel electrophoresis face an arduous task in maintaining a centralised and constantly evolving record of information relating to the characterisation of proteins and their responses following biological challenges. The Microbial Proteome Database (MPD) has been conceived as an in-house resource for complementing the plethora of genomic databases available for such organisms. The database utilises commercially available software to provide an electronic`lab book' of information obtained daily from 2-D electrophoresis gels, image analysis packages, protein characterisation methodologies, and biological experimentation. The MPD begins from a single 2-D gel image (a 2-D`reference map') with clickable spots that link to a`protein catalogue' (ProtCat) with spot information including protein identity, changes in expression determined under experimental conditions, cellular location, mass, and pI. The entry for each protein then contains further links to gel images corresponding to the presence of the particular protein within different subproteomes (as defined by the pH of narrow-and wide-range immobilised pH gradients or from differential extraction methods used to determine the location of the protein within a functional cell). The database currently contains information from strains of three microbial species (Escherichia coli, Pseudomonas aeruginosa and Staphylococcus aureus) and 32 master gel images. The rapid accessibility of information obtained from microbial proteomes is an essential step towards the integrated analysis of these organisms at the gene, transcript, protein and functional levels and will aid in reducing turnaround times between sample preparation and the discovery of molecules of biological significance.

Initial Proteome Analysis of Model MicroorganismHaemophilus influenzaeStrain Rd KW20

“…Currently, the most popular method employed to explore microbial proteomes is a combination of 2DE for separating and visualizing proteins and then MS for protein identification (7,8,12,13,19,60,61). The highest coverage for any proteome reported so far is a set of 502 proteins from H. influenzae (29% of all proteins predicted from the genome analysis), which was compiled from the results of multiple studies performed by several leading groups during a significant time period (34).…”

“…These situations can be addressed by a global postgenomic high-throughput approach commonly referred to as proteome analysis. Proteome analysis is aimed at studying all proteins expressed in a certain organism under certain conditions (1,5,7,8,18,27,34,35,37,38,53,60,61,(63)(64)(65)(66)(67). One of the central issues surrounding proteome analysis is accurately identifying proteins in complex mixtures extracted from the cells.…”

Perspectives for mass spectrometry and functional proteomics

“…There are now a large number of efforts to map all proteins of particular cell or tissue types on 2D gels. Within roughly the last year, publications that have described mapping of total proteins included work on Haemophilus influenzae (Langen et al, 2000), Saccharomyces cerevisiae (Futcher et al, 1999; Hodges et al, 1999; Perrot et al, 1999; Costanzo et al, 2000), Caenorhabditis elegans (Costanzo et al, 2000), Escherichia coli , Pseudomonas aeruginosa and Staphylococcus aureus (Cordwell et al, 1999), mycobacteria (Mollenkopf et al, 1999), HeLa cells (Shaw et al, 1999), human brain proteins from the parietal cortex lobe (Langen et al, 1999) and from the hippocampus (Edgar et al, 2000), mouse brain proteins (Gauss et al, 1999), rat heart proteins (Li et al, 1999b), human prostate proteins (Nelson et al, 2000), cancerous human bladder cells (Celis et al, 1999) and human breast cells (Page et al, 1999a). Some of the many other such studies in progress (Hoogland et al, 1999, 2000) are summarized at http://www.expasy.ch/ch2d/.…”

Status of genome projects for nonpathogenic bacteria and archaea

“…Matrix-assisted desorption/ionization-time of flight (MALDI-TOF) mass spectrometry enables high-throughput screening of protein samples derived from two-dimensional gel electrophoresis 15 and increases the practicality of whole-proteome analysis of sequenced organisms. Databases, such as The Microbial Proteome Database (MPD) 16 , are currently being developed to concentrate all the information being gleaned from two-dimensional gel studies. The database currently contains information from E. coli, Pseudomonas aeruginosa, and Staphylococcus aureus, as well as 32 master gel images.…”