ABSTRACT
The US Public Health Emergency Medical Countermeasures Enterprise convened subject matter experts at the 2010 HHS Burkholderia Workshop to develop consensus recommendations for postexposure prophylaxis against and treatment for Burkholderia pseudomallei and B. mallei infections, which cause melioidosis and glanders, respectively. Drugs recommended by consensus of the participants are ceftazidime or meropenem for initial intensive therapy, and trimethoprim/sulfamethoxazole or amoxicillin/clavulanic acid for eradication therapy. For postexposure prophylaxis, recommended drugs are trimethoprim/sulfamethoxazole or co-amoxiclav. To improve the timely diagnosis of melioidosis and glanders, further development and wide distribution of rapid diagnostic assays were also recommended. Standardized animal models and B. pseudomallei strains are needed for further development of therapeutic options. Training for laboratory technicians and physicians would facilitate better diagnosis and treatment options.
Subject(s)
Anti-Bacterial Agents/administration & dosage , Burkholderia mallei/pathogenicity , Burkholderia pseudomallei/pathogenicity , Glanders/prevention & control , Melioidosis/prevention & control , Post-Exposure Prophylaxis/methods , Amoxicillin-Potassium Clavulanate Combination/administration & dosage , Animals , Ceftazidime/administration & dosage , Disease Models, Animal , Disease Susceptibility , Glanders/diagnosis , Glanders/drug therapy , Humans , Melioidosis/diagnosis , Melioidosis/drug therapy , Meropenem , Risk Factors , Thienamycins/administration & dosage , Trimethoprim, Sulfamethoxazole Drug Combination/administration & dosageABSTRACT
Residual dipolar couplings (RDCs) have recently emerged as a new tool in nuclear magnetic resonance (NMR) with which to study macromolecular structure and function in a solution environment. RDCs are complementary to the more conventional use of NOEs to provide structural information. While NOEs are local-distance restraints, RDCs provide long-range orientational information. RDCs are now widely utilized in structure calculations. Increasingly, they are being used in novel applications to address complex issues in structural biology such as the accurate determination of the global structure of oligonucleotides and the relative orientation of protein domains. This review briefly describes the theory and methods for obtaining RDCs and then describes the range of biological applications where RDCs have been used.
Subject(s)
Algorithms , Crystallography/methods , DNA/chemistry , Magnetic Resonance Spectroscopy/methods , Proteins/chemistry , RNA/chemistry , Macromolecular Substances , Nucleic Acid Conformation , Protein Binding , Protein Folding , Protein Structure, Tertiary , SolutionsABSTRACT
Structures of intermediates and transition states in protein folding are usually characterized by amide hydrogen exchange and protein engineering methods and interpreted on the basis of the assumption that they have native-like conformations. We were able to stabilize and determine the high-resolution structure of a partially unfolded intermediate that exists after the rate-limiting step of a four-helix bundle protein, Rd-apocyt b(562), by multidimensional NMR methods. The intermediate has partial native-like secondary structure and backbone topology, consistent with our earlier native state hydrogen exchange results. However, non-native hydrophobic interactions exist throughout the structure. These and other results in the literature suggest that non-native hydrophobic interactions may occur generally in partially folded states. This can alter the interpretation of mutational protein engineering results in terms of native-like side chain interactions. In addition, since the intermediate exists after the rate-limiting step and Rd-apocyt b(562) folds very rapidly (k(f) approximately 10(4) s(-1)), these results suggest that non-native hydrophobic interactions, in the absence of topological misfolding, are repaired too rapidly to slow folding and cause the accumulation of folding intermediates. More generally, these results illustrate an approach for determining the high-resolution structure of folding intermediates.
Subject(s)
Protein Folding , Kinetics , Models, Molecular , Nuclear Magnetic Resonance, BiomolecularABSTRACT
Recent methods of aligning proteins which were developed in order to measure residual dipolar couplings (RDCs) in solution can also be used for additional applications such as measuring the 15N CSA in the form of chemical shift differences, Deltadelta. A new XPLOR-NIH module has been developed and implemented for NMR structure refinement using the 15N Deltadelta data as restraints. The results of this refinement are shown using the protein Bax. This method should be amenable to any protein which can be studied by NMR. An analysis comparing the structural information provided by NH RDCs and the 15N Deltadelta is included.
Subject(s)
Algorithms , Anisotropy , Models, Molecular , Nitrogen Isotopes/chemistry , Nuclear Magnetic Resonance, Biomolecular/methods , Proteins/chemistry , Proto-Oncogene Proteins c-bcl-2 , Computer Simulation , Hydrogen/chemistry , Hydrogen Bonding , Magnetics , Protein Conformation , Proto-Oncogene Proteins/chemistry , Quality Control , Reproducibility of Results , Sensitivity and Specificity , bcl-2-Associated X ProteinABSTRACT
An analysis of backbone hydrogen bonds has been performed on nine high-resolution protein X-ray crystal structures. Backbone hydrogen-bond geometry is compared in the context of X-ray crystal structure resolution. A strong correlation between the hydrogen-bond distance, R(HO), and the hydrogen-bond angle, theta(NHO), is observed when the X-ray crystal structure resolution is <1.00 A. Ab initio calculations were performed to substantiate these results. The angle and distance limits found in our correlation for the backbone hydrogen-bond geometry can be used to evaluate the quality of protein structures and for further NMR structure refinement.
