Search

Aspergillus fumigatus is a recognised human pathogen, especially in immunocompromised individuals. The availability of the annotated A. fumigatus genome sequence will significantly accelerate our understanding of this organism. However, limited information is available with respect to the A. fumigatus proteome. Here, both a direct proteomic approach (2D-PAGE and MALDI-MS) and a sub-proteomic strategy involving initial glutathione affinity chromatography have been deployed to identify 54 proteins from A. fumigatus primarily involved in energy metabolism and protein biosynthesis. Furthermore, two novel eukaryotic elongation factor proteins (eEF1Bc), termed ElfA and B have been identified and phylogenetically confirmed to belong to the eEF1Bc class of GST-like proteins. One of these proteins (ElfA) has been purified to homogeneity, identified as a monomeric enzyme (molecular mass = 20 kDa; pI = 5.9 and 6.5), and found to exhibit glutathione transferase activity specific activities (mean ± standard deviation, n = 3) of 3.13 ± 0.27 and 3.43 ± 1.0 lmol/min/mg, using CDNB and ethacrynic acid, respectively. Overall, these data highlight the importance of new approaches to dissect the proteome of, and elucidate novel functions within, A. fumigatus. 2006 Elsevier Inc. All rights reserved.

Publisher Summary Glutathione transferases play an important role in the biotransformation and detoxication of electrophilic xenobiotics. The occurrence of glutathione transferase in animal species is widespread. Several transferase isoenzymes are isolated from rat liver. Six basic transferase isoenzymes in rat hepatic cytosol are characterized as binary combinations of four protein subunits. The presence of glutathione transferase is not restricted to the liver but also is demonstrated in extrahepatic organs. However, in comparison with the liver most other organs show considerably lower activity. One exception is rat testis, which also shows high transferase activity. In contrast with liver, and most other organs, a major part of the glutathione transferase activity in rat testis is borne by an isoenzyme, glutathione transferase 6-6, with an acidic isoelectric point. The purification of this enzyme, which accounts for approximately 50% of the cytosolic glutathione transferase activity, is described in this chapter. In addition to this major acidic isoenzyme, smaller amounts of the basic species, glutathione transferases 2-2, 3-3, 3-4, and 4-4 are identified in testis.