X-ray Electron Density Maps Research Articles

Long Carbon-Carbon Bonding beyond 2 Å in Tris(9-fluorenylidene)methane.

We report on our investigation of C-C bonding longer than 2.0 Å, which can be realized by perpendicularly facing two fluorenyl rings in the title compound. A small orbital overlap between the distantly positioned carbon atoms is observed as a small concentration of electrons on the X-ray electron density map. The highest occupied molecular orbital (HOMO) and the lowest unoccupied molecular orbital (LUMO) of the compound originate from the in-phase and out-of-phase interactions of the overlapping orbitals, respectively, with a gap of 2.39 eV. Solid-state 13C NMR spectroscopy shows a sharp peak at 82.9 ppm for the long-bonded carbons, and a CASSCF(6,6) calculation indicates small diradical character. The experimental and theoretical analyses reveal sufficient covalent-bonding interaction in the long-bonded carbon pair.

Conformational Strain of Macrocyclic Peptides in Ligand-Receptor Complexes Based on Advanced Refinement of Bound-State Conformers.

Macrocyclic peptides are an important modality in drug discovery, but molecular design is limited due to the complexity of their conformational landscape. To better understand conformational propensities, global strain energies were estimated for 156 protein-macrocyclic peptide cocrystal structures. Unexpectedly large strain energies were observed when the bound-state conformations were modeled with positional restraints. Instead, low-energy conformer ensembles were generated using xGen that fit experimental X-ray electron density maps and gave reasonable strain energy estimates. The ensembles featured significant conformational adjustments while still fitting the electron density as well or better than the original coordinates. Strain estimates suggest the interaction energy in protein-ligand complexes can offset a greater amount of strain for macrocyclic peptides than for small molecules and non-peptidic macrocycles. Across all molecular classes, the approximate upper bound on global strain energies had the same relationship with molecular size, and bound-state ensembles from xGen yielded favorable binding energy estimates.

QFit-ligand reveals widespread conformational heterogeneity of drug-like molecules in X-ray electron-density maps

<i>qFit-ligand</i> reveals widespread conformational heterogeneity of drug-like molecules in X-ray electron-density maps

Adenovirus Composition, Proteolysis, and Disassembly Studied by In-depth Qualitative and Quantitative Proteomics

Using high-resolution MS-based proteomics in combination with multiple protease digestion, we profiled, with on average 90% sequence coverage, all 13 viral proteins present in an human adenovirus (HAdV) vector. This in-depth profile provided multiple peptide-based evidence on intrinsic protease activity affecting several HAdV proteins. Next, the generated peptide library was used to develop a targeted proteomics method using selected reaction monitoring (SRM) aimed at quantitative profiling of the stoichiometry of all 13 proteins present in the HAdV. We also used this method to probe the release of specific virus proteins initiated by thermal stimulation, mimicking the early stage of HAdV disassembly during entry into host cells. We confirmed the copy numbers of the most well characterized viral capsid components and established the copy numbers for proteins whose stoichiometry has so far not been accurately defined. We also found that heating HAdV induces the complete release of the penton base and fiber proteins as well as a substantial release of protein VIII and VI. For these latter proteins, maturational proteolysis by the adenoviral protease leads to the differential release of fragments with certain peptides being fully released and others largely retained in the AdV particles. This information is likely to be beneficial for the ongoing interpretation of high resolution cryoEM and x-ray electron density maps.

Structure-Based Prediction of the Mobility and Disorder of Water Molecules at Protein-DNA Interface

Water plays an invaluable role in governing the structure, stability, dynamics, and function of biomolecules, which has also been demonstrated to be critical in mediating biomolecular recognition and association. Accurate determination of the dynamic behavior of water molecules at biological complex interface is important for the understanding of the molecular mechanism of water contributing to the binding between biomolecules and could be exploited as an alternative tool to refine the water's positions in X-ray electron density map. In the present study, a method called local hydrophobic descriptors (LHDs) is used to characterize the hydrophobic landscapes of the hydration sites at protein-DNA interface. The resulting variables of the characterization are then correlated with the experimentally measured B-factor values of 4445 elaborately selected water samples derived from a panel of thematically nonredundant, high-quality protein-DNA interfaces by using a variety of machine learning methods, including PLS, BPNN, SVM, LSSVM, RF, and GP. The results show that the dynamic behavior of interfacial water molecules is primarily governed by the local hydrophobic feature of the hydration sites that water molecules located, and the nonlinear dependence dominates over the linear component in the water B-factor system. We expect that this structured-based approach can be used for predicting the B-factor profile of other biomolecules as well.

Structural analysis of compound I in hemoproteins: Study on Proteus mirabilis catalase

Ferryl catalysis has attracted considerable interest, because a diverse variety of enzymes use ferryl intermediates to perform difficult chemistry. The structure of the reactional intermediate compound I of Proteus mirabilis catalase (PMC) has been solved using time-resolved X-ray diffraction techniques and single crystal microspectrophotometry. Formation of compound I is characterized by significant changes in the absorbance spectrum, and the creation of an oxoferryl group on the distal side of the heme. This group is clearly visible in the X-ray electron density maps. An unidentified electron density, likely to be an anion because of the nature of its environment, appears during the reaction, in a site distant from the heme. The structure of compound I in PMC is compared with that of compound I in cytochrome c peroxidase (CCP).

A model-building procedure with particular application to proteins

A procedure for fitting a molecular model with fixed bond lengths and bond angles to a set of Cartesian coordinates has been developed. This procedure is designed to fit coordinates obtained from an X-ray electron-density map, although it can also fit coordinate sets from other sources. It differs from other model-building methods which use fixed bond lengths and bond angles in that it takes cognizance of certain information present in the electron-density map, which is ignored in other methods. The inherent uncertainty in the values of the coordinates determined from the map influences the fitting; i.e., if the computed location of an atom of the model is within the uncertainty assigned to the corresponding coordinate derived from the map, the fit of the atom of the model is assigned as exact. Also, the fitting process may begin anywhere along the molecular chain, in contrast to other procedures which commence fitting at or near one end of the chain. Thus, the present procedure can avoid errors that might arise if the starting point were chosen in a region where the atoms are poorly defined, as they often are at the ends of the chain. The procedure is designed to be used with a large amount of operator intervention, making it fairly flexible. Complete mathematical details of the method are given. A Fortran IV computer program using the method to fit a polypeptide model to a set of Cartesian coordinates has been written. The program has been used to fit a model of bovine pancreatic trypsin inhibitor to Cartesian coordinates derived from the 2.5 Å resolution electron-density map. The r.m.s. deviation between the model and the weighted coordinates from the map was 0.49 Å. As a preliminary step in a refinement of the 2.5 Å structure by potential-energy-constrained model building, the model obtained here was subjected to energy minimization with the atomic coordinates constrained to remain 'close' to the original guide points.

Three-fold structural pattern in the soybean trypsin inhibitor (Kunitz)

The molecule contains three very similar irregular Y-shaped lobes of antiparallel twisted β-sheet, which are grouped symmetrically round a central axis and linked by hydrogen bonds to form a six-stranded barrel. Each lobe can be superposed on either neighbour by a rotation of approximately 120 °. Of the 160 residues seen in the X-ray electron density map, 101 may be superposed onto other residues within a root-mean-square distance of 2.1 Å. The bond which reacts with trypsin lies on a loop between the first two lobes. It is suggested that the protein evolved from a primitive symmetrical trimer of identical subunits by tandem gene triplication.

Amino acid sequence of Paracoccus denitrificans cytochrome c550.

The amino acid sequence of Paracoccus (formerly Micrococcus) denitrificans cytochrome c550 has been established by a combination of standard chemical techniques and interpretation of a 2.5 A resolution x-ray electron density map. Peptides derived from a trypsin digest were chemically sequenced, and then ordered by fitting them to the density map. The amino acid compositions of chymotryptic peptides confirmed the x-ray map ordering the tryptic peptides. The amino acid sequence of this respiratory, prokaryotic cytochrome with 134 residues is discussed in relation to those of eukaryotic respiratory cytochrome c (103 to 113 amino acids), and prokaryotic, photosynthetic c2 (103 to 124 amino acids). At the primary structure level, c and c550 differ no more from cytochromes c2 than the various cytochromes c2 do from one another. It is suggested that the respiratory electron transport chain in prokaryotes and eukaryotes is a relatively late evolutionary offshoot of the photosynthetic electron transport chain in purple non-sulfur bacteria.