Current Proteomics - Volume 5, Issue 1, 2008

The advent of the post genomics era coupled with advances in proteomics presents unprecedented opportunities for expedited discovery of microbial targets for subsequent development of novel modalities for management of infectious diseases. Here, we review a new application of proteomics called Proteomics-based Expression Library Screening (PELS). This functional proteomics tool permits rapid definition of comprehensive microbial immunoproteomes (microbial proteins expressed in vivo that elicit and interact with the host humoral immune response), a subset of which have excellent potential as diagnostic, drug and vaccine targets. PELS is a platform technology that is applicable to any sequenced pathogen. It offers several advantages over existing methods for global identification of immunogenic microbial proteins, and is both a rapid alternative and an adjunct to emerging protein antigen array/chip technologies.

Protein-protein interactions are known to play significant roles in many biological processes. Knowing the detailed structure of protein-protein complex has become a very important issue in biological sciences. Protein-protein docking methods have been developed for many years in predicting the 3D-structure of protein-protein complexes when the structures of their protein components are available. The protein-protein docking problem involves two key elements: search algorithm and scoring function. The scoring function should be able to recognize the correctly docked complexes from incorrect ones and the search algorithm explores the huge conformational space extensively to find a nearly correct association of the protein components. A docking method becomes successful by combining the two key elements properly and adapting reasonable procedures. In this review, a brief background of protein-protein docking is introduced. Several widely-used docking programs are described and compared. The Critical Assessment of PRedicted Interactions (CAPRI) community-wide experiment, where docking methods have been tested over the past several years, is presented. In conclusion, future directions of protein-protein docking and its applications will be discussed.

Drug development, protein functions and structure elucidation can greatly benefit from the study of interactions between proteins and small molecules. The binding of a ligand to a protein can elicit a variety of molecular and cellular responses, and thus identifying a native ligand and understanding its interplay with a given protein can shed light about the protein function as well as its potential role as a drug target. Furthermore, the evaluation of drug interactions with protein targets is a widespread activity for efficient drug development. Mass Spectrometry (MS) can play a key role in the characterization of protein-ligand binding. For the detailed structural analysis of covalent-modified peptides, Ion Trap (IT), Time-Of-Flight (TOF) and hybrid mass spectrometers have been used as powerful tools. When the nature of the protein-ligand interaction is non-covalent, Electrospray Ionization (ESI) is the ionization technique of choice to gently and efficiently ionize and analyze these non-covalent complexes under nondenaturing conditions. Due to the fact that in crude biological samples ligand-unbound proteins are much more abundant than small molecule-bound proteins, it is difficult to identify the protein targets of small molecules in this type of samples. To address this problem, affinity-based approaches (e.g. affinity chromatography and immunoprecipitation) can be used to enrich for classes of proteins based on their affinity or activity toward specific ligands. As a complementary or alternative approach, the specific scanning modes of hybrid mass spectrometers allow ion filtering of ligand-bound molecules by selective analysis of small molecule-derived diagnostic fragment ions, and can help elucidate small molecule-protein binding sites. This review summarizes concepts and principles of protein separation approaches as well as mass spectrometric tools aimed at studying the interaction between small molecules and proteins. Selected examples from recently published work are described and discussed.

Healthcare of permanently growing cohort of diabetic patients is a serious economical problem in most industrialized countries including China and India. Diabetes mellitus (DM) frequently results in diverse severe complications, such as retinopathy, nephropathy, silent ischemia, dementia, and cancer. Oxidative damage to DNA is well documented for DM-patients. Individual sensitivity to oxidative stress varies among DM-patients and results in a cascade of chronic complications appearing as the “domino-effect”. Proliferative diabetic retinopathy is an early indicator for individual predisposition to chronic complications responsible for the majority of morbidity and mortality. In DM, changes in mitochondrial protein repertoire result in premature aging and plenty of pathologies including neurodegeneration and cancer. In diabetic care, much attention is focused on renal and urinary proteomics. However, the diagnosis of nephropathy can be reliably made only in patients with macroalbuminuria in the presence of diabetic retinopathy. Since 1/3-part of urinary proteome consists of plasma proteins and changes in plasma proteome occur up-stream towards chronic damage of organ systems, plasma proteomics possesses particular predictive power. Promising approach of a combination of plasma proteome and gene expression profiling in circulating leukocytes is currently discussed for development of potent diagnostic/ therapeutic targets. Estimation of gelatinase activity in serum combined with expression profiling of selected stress proteome genes in circulating leukocytes has been proposed for predictive imaging system of chronic DM-complications that allows initiation of appropriate therapy and consequent planning of personalized treatment. Expected long-term deliverables of nutri-proteomics is a nutrition individually optimized for prevention of chronic DM-complications.

3,5,3'-triiodo-L-thyronine (T3) modulates development and growth, and in adult life it exerts a profound effect on basal metabolic rate, increasing respiration rate and simultaneously lowering metabolic efficiency. It mainly acts through the coordinated and synergistic modulation of both nuclear and mitochondrial genome expressions giving rise to a complex network of factors and cellular events not yet completely defined. Thus, understanding the effects of T3 requires investigations at several levels [such as genes and gene transcripts (genome and transcriptome), proteins and metabolites, functions and metabolic assessment (proteome and metabolome)]. As yet, the ultimate effects of T3 on tissue-proteomes remain largely unknown. The proteins are excellent targets in disease diagnostics, prognostics, and therapeutics, and therefore proteomic approaches [such as two-dimensional gel electrophoresis (2D-E), two-dimensional liquid chromatography (2-DL), and mass spectrometry (MS)] represent powerful tools for a) the discovery of novel hormone/drug targets and biomarkers, and b) the study of in vivo and in vitro hormone effects on cellular metabolism and protein expressions. Increasingly proteomic techniques are being adopted, in particular to avoid the limitations inherent in the more classical approaches, to solve analytical problems and obtain a more comprehensive identification and characterization of molecular events associated with patho-physiological conditions. Here, we review the new leads emerging from the application of comparative proteomics to the actions of thyroid hormones, and we overview the technologies that can improve the resolution of proteins differing in hydrophobicity, intracellular location, complex formation, and membrane binding.

There is a high level of interest in polyunsaturated fatty acids (PUFAs) because of their purported health benefits, especially docosahexaenoic acid (DHA). But a high intake of them may increase the susceptibility to lipid peroxidation of a biological system. Lipid peroxidation has been implicated in the pathogenesis of numerous diseases including atherosclerosis, diabetes, cancer and aging. It is well established that the end-products of lipid peroxidation cause protein damage by means of reactions with lysine amino groups, cysteine sulfhydryl groups, and histidine imidazole groups. PUFAs in tissue are important sources for the formation of endogenous protein adducts, the relative contribution of different PUFAs in the formation of these protein adducts has been reviewed.