As a first step in establishing a proteome database for maize, we have embarked on the identification of the leaf proteins resolved on two-dimensional (2-D) gels. We detected nearly 900 spots on the gels with a pH 4-7 gradient and over 200 spots on the gels with a pH 6-11 gradient when the proteins were visualized with colloidal Coomassie blue. Peptide mass fingerprints for 300 protein spots were obtained with matrix assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometer and 149 protein spots were identified using the protein databases. We also searched the pdbEST databases to identify the leaf proteins and verified 66% of the protein spots that had been identified using the protein databases. Sixty-seven additional protein spots were identified from expressed sequence tags (ESTs). Many abundant leaf proteins are present in multiple spots. Functions of over 50% of the abundant leaf proteins are either unknown or hypothetical. Our results show that EST databases in conjunction with peptide mass fingerprints can be used for identifying proteins from organisms with incomplete genome sequence information.

Methylglyoxal is associated with a broad spectrum of biological effects, including cytostatic and cytotoxic activities. It is detoxified by the glyoxylase system or by its reduction to lactaldehyde by methylglyoxal reductase. We show that methylglyoxal reductase (NADPH-dependent) is encoded by GRE2 (YOL151w ). We associated this activity with its gene by partially purifying the enzyme and identifying by MALDI-TOF the proteins in candidate bands on SDS-PAGE gels whose relative intensities correlated with specific activity through three purification steps. The candidate proteins were then purified using a glutathione-S -transferase tag that was fused to them, and tested for methylglyoxal reductase activity. The advantage of this approach is that only modest protein purification is required. Our approach should be useful for identifying many of the genes that encode the metabolic pathway enzymes that have not been associated with a gene (about 275 in S. cerevisiae, by our estimate).

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To review mechanisms of regulation of expression and the functionality of two isoforms of translation elongation factor eEF1A in mammalian cells. Results. eEF1A1 and eEF1A2 proteins are regulated by post-translational modifications, protein-protein and protein-tRNA interactions as well as by controlling the amount of their mRNAs in human cells. Conclusions. EEF1A1 mRNA levels in cancer cells may depend on the allelic copy number while the level of EEF1A2 mRNA may be controlled by micro RNAs. eEF1A2 protein activity in different cellular processes may be determined, in part, by its increased affinity for tRNA and viral RNAs as compared to eEF1A1. eEF1A1 activity can be regulated by its increased susceptibility to post-translational modifications (PTM) and protein-protein interactions (PTI) as compared to eEF1A2.

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