Intrinsically disordered proteins (IDPs) or intrinsically disordered regions (IDRs) is a class of biologically important proteins exhibiting specific biophysical characteristics. They lack a hydrophobic core, and their conformational behavior is strongly influenced by electrostatic interactions. IDPs and IDRs are highly dynamic, and a characterization of the motions of IDPs and IDRs is essential for their physically correct description. NMR together with molecular dynamics simulations are the methods best suited to such a task because they provide information about dynamics of proteins with atomistic resolution. Here, we present a study of motions of a disordered C-terminal domain of the delta subunit of RNA polymerase from Bacillus subtilis. Positively and negatively charged residues in the studied domain form transient electrostatic contacts critical for the biological function. Our study is focused on investigation of ps-ns dynamics of backbone of the delta subunit based on analysis of amide 15N NMR relaxation data and molecular dynamics simulations. In order to extend an informational content of NMR data to lower frequencies, which are more sensitive to slower motions, we combined standard (high-field) NMR relaxation experiments with high-resolution relaxometry. Altogether, we collected data reporting the relaxation at 12 different magnetic fields, resulting in an unprecedented data set. Our results document that the analysis of such data provides a consistent description of dynamics and confirms the validity of so far used protocols of the analysis of dynamics of IDPs also for a partially folded protein. In addition, the potential to access detailed description of motions at the timescale of tens of ns with the help of relaxometry data is discussed. Interestingly, in our case, it appears to be mostly relevant for a region involved in the formation of temporary contacts within the disordered region, which was previously proven to be biologically important.

• MeSH
• Amides MeSH
• DNA-Directed RNA Polymerases chemistry   MeSH
• Protein Conformation MeSH
• Magnetic Resonance Spectroscopy MeSH
• Molecular Dynamics Simulation MeSH
• Intrinsically Disordered Proteins * chemistry   MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Amides MeSH
• DNA-Directed RNA Polymerases MeSH
• Intrinsically Disordered Proteins * MeSH

Intrinsically disordered proteins are ubiquitous throughout all known proteomes, playing essential roles in all aspects of cellular and extracellular biochemistry. To understand their function, it is necessary to determine their structural and dynamic behavior and to describe the physical chemistry of their interaction trajectories. Nuclear magnetic resonance is perfectly adapted to this task, providing ensemble averaged structural and dynamic parameters that report on each assigned resonance in the molecule, unveiling otherwise inaccessible insight into the reaction kinetics and thermodynamics that are essential for function. In this review, we describe recent applications of NMR-based approaches to understanding the conformational energy landscape, the nature and time scales of local and long-range dynamics and how they depend on the environment, even in the cell. Finally, we illustrate the ability of NMR to uncover the mechanistic basis of functional disordered molecular assemblies that are important for human health.

• MeSH
• Protein Conformation MeSH
• Humans MeSH
• Magnetic Resonance Spectroscopy MeSH
• Nuclear Magnetic Resonance, Biomolecular MeSH
• Thermodynamics MeSH
• Intrinsically Disordered Proteins * chemistry   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Review MeSH
• Names of Substances
• Intrinsically Disordered Proteins * MeSH

Improving our understanding of nanosecond motions in disordered proteins requires the enhanced sampling of the spectral density function obtained from relaxation at low magnetic fields. High-resolution relaxometry and two-field NMR measurements of relaxation have, so far, only been based on the recording of one- or two-dimensional spectra, which provide insufficient resolution for challenging disordered proteins. Here, we introduce a 3D-HNCO-based two-field NMR experiment for measurements of protein backbone 15 N amide longitudinal relaxation rates. The experiment provides accurate longitudinal relaxation rates at low field (0.33 T in our case) preserving the resolution and sensitivity typical for high-field NMR spectroscopy. Radiofrequency pulses applied on six different radiofrequency channels are used to manipulate the spin system at both fields. The experiment was demonstrated on the C-terminal domain of δ subunit of RNA polymerase from Bacillus subtilis, a protein with highly repetitive amino-acid sequence and very low dispersion of backbone chemical shifts.

• Keywords
• Dynamics, High-resolution relaxometry, Intrinsically disordered proteins, Non-uniform sampling, Nuclear magnetic resonance, Relaxation,
• MeSH
• Bacillus subtilis enzymology   MeSH
• Bacterial Proteins chemistry   MeSH
• DNA-Directed RNA Polymerases chemistry   MeSH
• Nuclear Magnetic Resonance, Biomolecular * MeSH
• Recombinant Proteins chemistry   MeSH
• Intrinsically Disordered Proteins chemistry   MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Bacterial Proteins MeSH
• DNA-Directed RNA Polymerases MeSH
• Recombinant Proteins MeSH
• Intrinsically Disordered Proteins MeSH

Microtubule-associated protein 2c (MAP2c) is a 49-kDa intrinsically disordered protein regulating the dynamics of microtubules in developing neurons. MAP2c differs from its sequence homologue Tau in the pattern and kinetics of phosphorylation by cAMP-dependent protein kinase (PKA). Moreover, the mechanisms through which MAP2c interacts with its binding partners and the conformational changes and dynamics associated with these interactions remain unclear. Here, we used NMR relaxation and paramagnetic relaxation enhancement techniques to determine the dynamics and long-range interactions within MAP2c. The relaxation rates revealed large differences in flexibility of individual regions of MAP2c, with the lowest flexibility observed in the known and proposed binding sites. Quantitative conformational analyses of chemical shifts, small-angle X-ray scattering (SAXS), and paramagnetic relaxation enhancement measurements disclosed that MAP2c regions interacting with important protein partners, including Fyn tyrosine kinase, plectin, and PKA, adopt specific conformations. High populations of polyproline II and α-helices were found in Fyn- and plectin-binding sites of MAP2c, respectively. The region binding the regulatory subunit of PKA consists of two helical motifs bridged by a more extended conformation. Of note, although MAP2c and Tau did not differ substantially in their conformations in regions of high sequence identity, we found that they differ significantly in long-range interactions, dynamics, and local conformation motifs in their N-terminal domains. These results highlight that the N-terminal regions of MAP2c provide important specificity to its regulatory roles and indicate a close relationship between MAP2c's biological functions and conformational behavior.

• Keywords
• NMR relaxation, Tau protein (Tau), microtubule-associated protein (MAP), nuclear magnetic resonance (NMR), paramagnetic relaxation enhancement (PRE), protein conformation, small-angle X-ray scattering (SAXS),
• MeSH
• X-Ray Diffraction MeSH
• Phosphorylation MeSH
• Protein Conformation * MeSH
• Humans MeSH
• Scattering, Small Angle MeSH
• Plectin chemistry   metabolism   MeSH
• Microtubule-Associated Proteins chemistry   metabolism   MeSH
• src Homology Domains MeSH
• Protein Binding MeSH
• Binding Sites MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• MAP2 protein, human MeSH Browser
• PLEC protein, human MeSH Browser
• Plectin MeSH
• Microtubule-Associated Proteins MeSH

Description of protein dynamics is known to be essential in understanding their function. Studies based on a well established [Formula: see text] NMR relaxation methodology have been applied to a large number of systems. However, the low dispersion of [Formula: see text] chemical shifts very often observed within intrinsically disordered proteins complicates utilization of standard 2D HN correlated spectra because a limited number of amino acids can be characterized. Here we present a suite of triple resonance HNCO-type NMR experiments for measurements of five [Formula: see text] relaxation parameters ([Formula: see text], [Formula: see text], NOE, cross-correlated relaxation rates [Formula: see text] and [Formula: see text]) in doubly [Formula: see text],[Formula: see text]-labeled proteins. We show that the third spectral dimension combined with non-uniform sampling provides relaxation rates for almost all residues of a protein with extremely poor chemical shift dispersion, the C terminal domain of [Formula: see text]-subunit of RNA polymerase from Bacillus subtilis. Comparison with data obtained using a sample labeled by [Formula: see text] only showed that the presence of [Formula: see text] has a negligible effect on [Formula: see text], [Formula: see text], and on the cross-relaxation rate (calculated from NOE and [Formula: see text]), and that these relaxation rates can be used to calculate accurate spectral density values. Partially [Formula: see text]-labeled sample was used to test if the observed increase of [Formula: see text] [Formula: see text] in the presence of [Formula: see text] corresponds to the [Formula: see text] dipole-dipole interactions in the [Formula: see text],[Formula: see text]-labeled sample.

• Keywords
• Intrinsically disordered proteins, Non-uniform sampling, Nuclear magnetic resonance, Relaxation,
• MeSH
• Bacillus subtilis enzymology   MeSH
• DNA-Directed RNA Polymerases chemistry   MeSH
• Nitrogen Isotopes MeSH
• Carbon Isotopes MeSH
• Nuclear Magnetic Resonance, Biomolecular methods   MeSH
• Intrinsically Disordered Proteins chemistry   MeSH
• Hydrogen MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Carbon-13 MeSH Browser
• DNA-Directed RNA Polymerases MeSH
• Nitrogen Isotopes MeSH
• Carbon Isotopes MeSH
• Nitrogen-15 MeSH Browser
• Intrinsically Disordered Proteins MeSH
• Hydrogen MeSH

The Eighth Central European Conference "Chemistry towards Biology" was held in Brno, Czech Republic, on August 28-September 1, 2016 to bring together experts in biology, chemistry and design of bioactive compounds; promote the exchange of scientific results, methods and ideas; and encourage cooperation between researchers from all over the world. The topics of the conference covered "Chemistry towards Biology", meaning that the event welcomed chemists working on biology-related problems, biologists using chemical methods, and students and other researchers of the respective areas that fall within the common scope of chemistry and biology. The authors of this manuscript are plenary speakers and other participants of the symposium and members of their research teams. The following summary highlights the major points/topics of the meeting.

• Keywords
• ADME, drug delivery systems, biological chemistry, biomaterials, chemical biology, drug design, nanoparticles, natural compounds, proteins and nucleic acids, synthesis, targeting,
• MeSH
• Epigenesis, Genetic MeSH
• Chemistry, Pharmaceutical methods   MeSH
• Drug Delivery Systems MeSH
• Proteins chemistry   MeSH
• Drug Design MeSH
• Systems Biology MeSH
• Structure-Activity Relationship MeSH
• Publication type
• Congress MeSH
• Names of Substances
• Proteins MeSH

Adiabatically swept pulses were originally designed for the purpose of broadband spin inversion. Later, unexpected advantages of their utilization were also found in other applications, such as refocusing to excite spin echoes, studies of chemical exchange or fragment-based drug design. Here, we present new experiments to characterize fast (ps-ns) protein dynamics, which benefit from little-known properties of adiabatic pulses. We developed a strategy for measuring cross-correlated cross-relaxation (CCCR) rates during adiabatic pulses. This experiment provides a linear combination of longitudinal and transverse CCCR rates, which is offset-independent across a typical amide (15)N spectrum. The pulse sequence can be recast to provide accurate transverse CCCR rates weighted by the populations of exchanging states. Sensitivity can be improved in systems in slow exchange. Finally, the experiments can be easily modified to yield residue-specific correlation times. The average correlation time of motions can be determined with a single experiment while at least two different experiments had to be recorded until now.

• Keywords
• Adiabatic sweep, Cross-correlated cross relaxation, NMR, Protein dynamics,
• MeSH
• Protein Interaction Domains and Motifs MeSH
• Humans MeSH
• Nuclear Magnetic Resonance, Biomolecular methods   MeSH
• CREB-Binding Protein chemistry   MeSH
• Ubiquitin chemistry   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• CREBBP protein, human MeSH Browser
• CREB-Binding Protein MeSH
• Ubiquitin MeSH

Merozoite surface protein 2 (MSP2) of Plasmodium falciparum is an abundant, intrinsically disordered protein that is GPI-anchored to the surface of the invasive blood stage of the malaria parasite. Recombinant MSP2 has been trialled as a component of a malaria vaccine, and is one of several disordered proteins that are candidates for inclusion in vaccines for malaria and other diseases. Nonetheless, little is known about the implications of protein disorder for the development of an effective antibody response. We have therefore undertaken a detailed analysis of the conformational dynamics of the two allelic forms of MSP2 (3D7 and FC27) using NMR spectroscopy. Chemical shifts and NMR relaxation data indicate that conformational and dynamic properties of the N- and C-terminal conserved regions in the two forms of MSP2 are essentially identical, but significant variation exists between and within the central variable regions. We observe a strong relationship between the conformational dynamics and the antigenicity of MSP2, as assessed with antisera to recombinant MSP2. Regions of increased conformational order in MSP2, including those in the conserved regions, are more strongly antigenic, while the most flexible regions are minimally antigenic. This suggests that modifications that increase conformational order may offer a means to tune the antigenicity of MSP2 and other disordered antigens, with implications for vaccine design.

• MeSH
• Antigens, Protozoan chemistry   immunology   MeSH
• Protein Conformation MeSH
• Nuclear Magnetic Resonance, Biomolecular MeSH
• Plasmodium falciparum immunology   MeSH
• Protozoan Proteins chemistry   immunology   MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Antigens, Protozoan MeSH
• merozoite surface protein 2, Plasmodium MeSH Browser
• Protozoan Proteins MeSH