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Binding Surface (binding + surface)
Selected AbstractsIdentification of crucial residues for the antibacterial activity of the proline-rich peptide, pyrrhocoricinFEBS JOURNAL, Issue 17 2002Goran Kragol Members of the proline-rich antibacterial peptide family, pyrrhocoricin, apidaecin and drosocin appear to kill responsive bacterial species by binding to the multihelical lid region of the bacterial DnaK protein. Pyrrhocoricin, the most potent among these peptides, is nontoxic to healthy mice, and can protect these animals from bacterial challenge. A structure,antibacterial activity study of pyrrhocoricin against Escherichia coli and Agrobacterium tumefaciens identified the N-terminal half, residues 2,10, the region responsible for inhibition of the ATPase activity, as the fragment that contains the active segment. While fluorescein-labeled versions of the native peptides entered E. coli cells, deletion of the C-terminal half of pyrrhocoricin significantly reduced the peptide's ability to enter bacterial or mammalian cells. These findings highlighted pyrrhocoricin's suitability for combating intracellular pathogens and raised the possibility that the proline-rich antibacterial peptides can deliver drug leads into mammalian cells. By observing strong relationships between the binding to a synthetic fragment of the target protein and antibacterial activities of pyrrhocoricin analogs modified at strategic positions, we further verified that DnaK was the bacterial target macromolecule. Inaddition, the antimicrobial activity spectrum of native pyrrhocoricin against 11 bacterial and fungal strains and the binding of labeled pyrrhocoricin to synthetic DnaK D-E helix fragments of the appropriate species could be correlated. Mutational analysis on a synthetic E. coli DnaK fragment identified a possible binding surface for pyrrhocoricin. [source] The crystal structure of microtubule-associated protein light chain 3, a mammalian homologue of Saccharomyces cerevisiae Atg8GENES TO CELLS, Issue 7 2004Kenji Sugawara Microtubule-associated protein light chain 3 (LC3), a mammalian homologue of yeast Atg8, plays an essential role in autophagy, which is involved in the bulk degradation of cytoplasmic components by the lysosomal system. Here, we report the crystal structure of LC3 at 2.05 Å resolution with an R-factor of 21.8% and a free R-factor of 24.9%. The structure of LC3, which is similar to those of Golgi-associated ATPase enhancer of 16 kDa (GATE-16) and GABAA receptor-associated protein (GABARAP), contains a ubiquitin core with two , helices, ,1 and ,2, attached at its N-terminus. Some common and distinct features are observed among these proteins, including the conservation of residues required to form an interaction among ,1, ,2 and the ubiquitin core. However, the electrostatic potential surfaces of these helices differ, implicating particular roles to select specific binding partners. Hydrophobic patches on the ubiquitin core of LC3, GABARAP and GATE-16 are well conserved and are similar to the E1 binding surface of ubiquitin and NEDD8. Therefore, we propose that the hydrophobic patch is a binding surface for mammalian Atg7 similar to a ubiquitin-like conjugation system. We also propose the functional implications of the ubiquitin fold as a recognition module of target proteins. [source] Communication between E,54, promoter DNA and the conserved threonine residue in the GAFTGA motif of the PspF ,54 -dependent activator during transcription activationMOLECULAR MICROBIOLOGY, Issue 2 2004Patricia Bordes Summary Conversion of E,54 closed promoter complexes to open promoter complexes requires specialized activators which are members of the AAA (ATPases Associated with various cellular Activities) protein family. The ATP binding and hydrolysis activity of E,54 activators is used in an energy coupling reaction to remodel the E,54 closed promoter complex and to overcome the ,54 -imposed block on open complex formation. The remodelling target for the AAA activator within the E,54 closed complex includes a complex interface contributed to by Region I of ,54, core RNA polymerase and a promoter DNA fork junction structure, comprising the E,54 regulatory centre. One ,54 binding surface on E,54 activators is a conserved sequence known as the GAFTGA motif. Here, we present a detailed characterization of the interaction between Region I of ,54 and the Escherichia coli AAA ,54 activator Phage shock protein F. Using E,54 promoter complexes that mimic different conformations adopted by the DNA during open complex formation, we investigated the contribution of the conserved threonine residue in the GAFTGA motif to transcription activation. Our results suggest that the organization of the E,54 regulatory centre, and in particular the conformation adopted by the ,54 Region I and the DNA fork junction structure during open complex formation, is communicated to the AAA activator via the conserved T residue of the GAFTGA motif. [source] Characterization of the binding surface of the translocated intimin receptor, an essential protein for EPEC and EHEC cell adhesionPROTEIN SCIENCE, Issue 12 2007Nathan T. Ross Abstract The translocated intimin receptor (TIR) of enteropathogenic and enterohemorrhagic Escherichia coli (EPEC and EHEC) is required for EPEC and EHEC infections, which cause widespread illness across the globe. TIR is translocated via a type-III secretion system into the intestinal epithelial cell membrane, where it serves as an anchor for E. coli attachment via its binding partner intimin. While many aspects of EPEC and EHEC infection are now well understood, the importance of the intermolecular contacts made between intimin and TIR have not been thoroughly investigated. Herein we report site-directed mutagenesis studies on the intimin-binding domain of EPEC TIR, and how these mutations affect TIR-intimin association, as analyzed by isothermal titration calorimetry and circular dichroism. These results show how two factors govern TIR's binding to intimin: A three-residue TIR hot spot is identified that largely mediates the interaction, and mutants that alter the ,-hairpin structure of TIR severely diminish binding affinity. In addition, peptides incorporating key TIR residues identified by mutagenesis are incapable of binding intimin. These results indicate that hot spot residues and structural orientation/preorganization are required for EPEC, and likely EHEC, TIR-intimin binding. [source] Mimicry of dimerization by synthetic peptides designed to target homologous regions of proteinsPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 3 2003Donard S. Dwyer Abstract Rapid progress in sequencing various genomes has highlighted the need for the development of biochemical reagents for the detection of thousands of expressed gene products. The magnitude of this detection problem exceeds current technical capabilities. In an attempt to address this shortcoming, a novel approach has been developed called mimicry of dimerization. Peptide tags have been designed to bind to a specific region of parvalbumin on the basis of amino acid sequence homology with this segment. Multivalent ligands were produced by coupling the synthetic peptides to activated dextran polymers and binding was assessed by chemiluminescence of enhanced avidity reactions using a high density of target protein at the binding surface. Binding of the peptide ligands to parvalbumin was strongest under assay conditions that enriched for native monomeric protein and was affected by pH, temperature and solvent conditions. The results suggest that it should be possible to develop specific reagents for tagging proteins on the basis of sequence and secondary structure information. [source] | |||||||||||||