Protein-peptide interactions/ complexes

Although issues thus remain with simpler aspects of protein/peptide interactions with phospholipid membranes, there has been a clear shift in the past few years to more complex membranes, notably those containing key nonlipid membrane components such as LPS, Upoteichoic acid, and proteoglycans. Protein/peptide interactions with such LPSs are important in various biological contexts, including Upoprotein deposition at proteoglycan-covered endothelial surfaces in atherosclerosis, lectin... 

Figure 5.11. Generic approaches to identify interacting proteins within complexes. The complex is isolated from cells by affinity purification using a tag sequence attached to a protein known to be in the complex. Alternatively, the complex can be immunprecipitated with an antibody to one of the proteins in the complex. The proteins are resolved by polyacrylamide gel electrophoresis, proteolyzed, and the mass of the resulting peptides is determined by mass spectrometry. Alternatively, the proteins can be proteolyzed and the resulting peptides resolved by liquid chromatography. The peptide masses are then determined by mass spectrometry and used for database searching to identify the component proteins.

The general types of protein-protein interactions that occur in cells include receptor-ligand, enzyme-substrate, multimeric complex formations, structural scaffolds, and chaperones. However, proteins interact with more targets than just other proteins. Protein interactions can include protein-protein or protein-peptide, protein-DNA/RNA or protein-nucleic acid, protein-glycan or protein-carbohydrate, protein-lipid or protein-membrane, and protein-small molecule or protein-ligand. It is likely that every molecule within a cell has some kind of specific interaction with a protein. 

The use of PIR compounds to study protein interactions is a significant advance over the use of standard homobifunctional crosslinkers. The unique design of the PIR reagent facilitates deconvolution of putative protein interaction complexes through a simplified mass spec analysis. The software can ignore all irrelevant peak data and just focus analysis on the two labeled peptide peaks, which accompany the reporter signal of appropriate mass. This greatly simplifies the bioinformatics of data analysis and provides definitive conformation of protein-protein crosslinks. 

With this background there is an obvious call for novel strategies to follow changes of complex molecular patterns of different stress-related diseases over days, weeks, months, and years as an effect of lifestyle and the psychosocial environment to reflect the effects of unhealthy environments. The molecular interactions between the brain and the immune system in health and disease are reflected in the circulatory system as the white blood cells, the lymphocytes, mimic ongoing activities in the brain. By using lymphocytes from patients with psychosomatic-psychiatric diseases we can find detailed information about protein-peptide translational modifications and transformation essential for the development of new approaches that can prevent and treat major psychiatric diseases. 

While these complex model heme proteins have a large potential for functionalization, an interesting approach that is very different has been taken by other workers in that the heme itself functions as the template in the formation of folded peptides. In these models peptide-peptide interactions are minimized and the driving force for folding appears to be the interactions between porphyrin and the hydrophobic faces of the amphiphiUc peptides. The amino acid sequences are too small to permit peptide-peptide contacts as they are separated by the tetrapyrrole residue. These peptide heme conjugates show well-re-solved NMR spectra and thus well-defined folds and the relationship between structure and function can probably be determined in great detail when functions have been demonstrated [22,23,77]. They are therefore important model systems that complement the more complex proteins described above. 

These minimalistic peptide scaffolds potentially provide a biologically relevant laboratory in which to explore the details of heme-peptide interactions and, with development, perhaps approach the observed range of natural heme protein fimction. These heme-peptide systems are more complex than typical small molecule bioinorganic porphyrin model compoimds, and yet are seemingly not as enigmatic as even the smallest natural heme proteins. Thus, in the continuum of heme protein model complexes these heme-peptide systems lie closer to, but certainly not at, the small molecule limit which allows for the effects of single amino acid changes to be directly elucidated. 

Protein-Ligand Interactions Exchange processes and determination of ligand conformation and protein-ligand contacts, 238, 657 nuclear magnetic studies of protein-peptide complexes,... 

In the following, some examples will be discussed to highlight the scope and the limitations of ACE for the detection of complex formation between DNA samples and interaction partners such as proteins, peptides, DNA or analogs, and smaller molecules. 

The possibility to carry out conformational studies of peptides at low concentrations and in the presence of complex biological systems represents a major advantage of fluorescence spectroscopy over other techniques. Fluorescence quantum yield or lifetime determinations, anisotropy measurements and singlet-singlet resonance energy transfer experiments can be used to study the interaction of peptides with lipid micelles, membranes, proteins, or receptors. These fluorescence techniques can be used to determine binding parameters and to elucidate conformational aspects of the interaction of the peptide with a particular macro-molecular system. The limited scope of this chapter does not permit a comprehensive review of the numerous studies of this kind that have been carried and only a few general aspects are briefly discussed here. Fluorescence studies of peptide interactions with macromolecular systems published prior to 1984 have been reviewed. 

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