Simulation of time-resolved vibronic spectra and the possibility of analyzing molecules with similar spectral properties

The possibility of using time-resolved vibronic spectroscopy for spectral analysis of mixtures of chemical compounds with similar optical properties, when traditional methods (based on stedy-state spectra) are inefficient, is demonstrated by using the method of computer simulation. The analysis is carried out by the example of molecules of a series of polyenes (butadiene, hexadiene, octatetraene, decapentaene, and decatetraene), their various cis-and trans-rotational isomers, and phenyl-substituted polyenes. Ranges of relative concentrations of molecules similar in their spectral properties, where reliable interpretation of time-resolved spectra of mixtures and both qualitative and quantitative analyses are possible, are determined. The use of computer simulation methods for optimizing full-scale experiments in femtosecond spectroscopy is shown to hold much promise.     

Ab initio calculations of vibronic activity in phosphorescence microwave double resonance spectra of p-dichlorobenzene

The phosphorescence spectrum of p-dichlorobenzene has been calculated using multiconfiguration self-consistent-field wave functions and the quadratic response technique. Attention has been paid to the intensity distribution of the singlet–triplet (³B_1u¹A_g) transition through a number of vibronic subbands. The second order spin–orbit coupling (SOC) contribution to the spin splitting of the ³B_1u (^3*) state is found to be almost negligible, and the calculations therefore provide a good estimate for the zero-field splitting (ZFS) parameters based only on the electron spin–spin coupling expectation values. Nuclear quadrupole resonance constants for the different Cl isotopes are also calculated to accomplish the ZFS assignment. The electric dipole activity of the spin sublevels in the triplet–singlet transitions to the ground-state vibrational levels is estimated by calculations of derivatives using distorted geometries which are shifted from the equilibrium position along different vibrational modes. A vibrational analysis of the phosphorescence spectrum, based on the SOC-induced mixing of the singlet and triplet states calculated along different vibrational modes, provides reasonable agreement with experimental data.Acknowledgment O. R.-P. would like to thank the European MOLPROP network for support. The authors thank Alexander Baev for fruitful discussions. This work was supported by the Swedish Royal Academy of Science (KVA).     

Crystal and Molecular Structure of the Nitramino Derivatives of Tetrazole and 1,2,4-Triazole. III. 5-Nitraminotetrazole Lithium Salt Monohydrate

The structure of 5-nitraminotetrazole lithium salt monohydrate was determined by X-ray diffraction analysis. Crystals are monoclinic, space group P2₁/c; a = 8.3789(5), b = 10.1872(6), c = 6.6709(5) ; = 106.63(1)°; V = 545.60(98) ³; Z = 4; _calc = 1.875 g/cm³. The anion has a planar nitrimine structure with a delocalized negative charge. Each lithium cation (c.n. 5) is bound to three anions and two hydration water molecules. Both oxygen atoms of the nitro groups and the N(3) atom of the tetrazole ring are involved in cation coordination. The geometrical characteristics of the anion are similar to those found for other monosalts of 5-nitraminotetrazole.     

Visual detection of cysteine and homocysteine

The determination of cysteine and homocysteine levels is of great current interest for the monitoring of desease states. A new colorimetric method for the simultaneous detection of l-cysteine and l-homocysteine has been developed. A fluorescein derivative reacts with the above amino acids, producing their respective thiazolidines resulting in color changes. Interference from other amino acids and proteins is minimal.     

Automated linkage of proteins and payloads producing monodisperse conjugates

Controlled protein functionalization holds great promise for a wide variety of applications. However, despite intensive research, the stoichiometry of the functionalization reaction remains difficult to control due to the inherent stochasticity of the conjugation process. Classical approaches that exploit peculiar structural features of specific protein substrates, or introduce reactive handles via mutagenesis, are by essence limited in scope or require substantial protein reengineering. We herein present equimolar native chemical tagging (ENACT), which precisely controls the stoichiometry of inherently random conjugation reactions by combining iterative low-conversion chemical modification, process automation, and bioorthogonal trans-tagging. We discuss the broad applicability of this conjugation process to a variety of protein substrates and payloads.

Controlled protein functionalization holds great promise for a wide variety of applications.

Applications of protein conjugates are limitless, including imaging, diagnostics, drug delivery, and sensing.^1–4 In many of these applications, it is crucial that the conjugates are homogeneous.⁵ The site-selectivity of the conjugation process and the number of functional labels per biomolecule, known as the degree of conjugation (DoC), are crucial parameters that define the composition of the obtained products and are often the limiting factors to achieving adequate performance of the conjugates. For instance, immuno-PCR, an extremely sensitive detection technique, requires rigorous control of the average number of oligonucleotide labels per biomolecule (its DoC) in order to achieve high sensitivity.⁶ In optical imaging, the performance of many super-resolution microscopy techniques is directly defined by the DoC of fluorescent tags.⁷ For therapeutics, an even more striking example is provided by antibody–drug conjugates, which are prescribed for the treatment of an increasing range of cancer indications.⁸ A growing body of evidence from clinical trials indicates that bioconjugation parameters, DoC and DoC distribution, directly influence the therapeutic index of these targeted agents and hence must be tightly controlled.⁹Standard bioconjugation techniques, which rely on nucleophile–electrophile reactions, result in a broad distribution of different DoC species (Fig. 1a), which have different biophysical parameters, and consequently different functional properties.¹⁰Open in a separate windowFig. 1Schematic representation of the types of protein conjugates.To address this key issue and achieve better DoC selectivity, a number of site-specific conjugation approaches have been developed (Fig. 1b). These techniques rely on protein engineering for the introduction of specific motifs (e.g., free cysteines,¹¹ selenocysteines,¹² non-natural amino acids,^13,14 peptide tags recognized by specific enzymes^15,16) with distinct reactivity compared to the reactivity of the amino acids present in the native protein. These motifs are used to simultaneously control the DoC (via chemo-selective reactions) and the site of payload attachment. Both parameters are known to influence the biological and biophysical parameters of the conjugates,¹¹ but so far there has been no way of evaluating their impact separately.The influence of DoC is more straightforward, with a lower DoC allowing the minimization of the influence of payload conjugation on the properties of the protein substrate. The lowest DoC that can be achieved for an individual conjugate is 1 (corresponding to one payload attached per biomolecule). It is noteworthy that DoC 1 is often difficult to achieve through site-specific conjugation techniques due to the symmetry of many protein substrates (e.g., antibodies). Site selection is a more intricate process, which usually relies on a systematic screening of conjugation sites for some specific criteria, such as stability or reactivity.¹⁷Herein, we introduce a method of accessing an entirely new class of protein conjugates with multiple conjugation sites but strictly homogenous DoCs (Fig. 1c). To achieve this, we combined (a) iterative low conversion chemical modification, (b) process automation, and (c) bioorthogonal trans-tagging in one workflow.The method has been exemplified for protein substrates, but it is applicable to virtually any native bio-macromolecule and payload. Importantly, this method allows for the first time the disentangling of the effects of homogeneous DoC and site-specificity on conjugate properties, which is especially intriguing in the light of recent publications revealing the complexity of the interplay between payload conjugation sites and DoC for in vivo efficacy of therapeutic bioconjugates.¹⁸ Finally, it is noteworthy that this method can be readily combined with an emerging class of site-selective bioconjugation reagents to produce site-specific DoC 1 conjugates, thus further expanding their potential for biotechnology applications.¹⁹     

Crystal and molecular structure of nitraminotetrazole and nitramino-1,2,4-triazole. IV. 5-nitraminotetrazole sodium salt sesquihydrate

The structure of 5-nitraminotetrazole sodium salt sesquihydrate was determined by X-ray diffraction. The crystals are monoclinic, space group P2₁/c;a = 3.551(1) Å, b = 21.834(4) Å, c = 9.075(2) Å; = 110.68(3)°; V = 658.3(2) ų; Z = 4; _calc = 1.807 g/cm³. The anion is planar and has an intramolecular hydrogen bond. The negative charge of the anion is localized on one of the oxygens of the nitro group. The sodium cation (c.n.6) is coordinated by three oxygen atoms of different anions and three oxygens of crystallization water. One of the crystallization water molecules is disordered in the unit cell. The anions are hydrogen-bonded with each other and with crystallization water molecules.Original Russian Text Copyright © 2004 by A. M. Astakhov, A. D. Vasiliev, M. S. Molokeev, L. A. Kruglyakova, A. M. Sirotinin, and R. S. StepanovTranslated from Zhurnal Strukturnoi Khimii, Vol. 45, No. 3, pp. 562–565, May–June 2004.     

Generation of new landomycins by combinatorial biosynthetic manipulation of the LndGT4 gene of the landomycin E cluster in S. globisporus

A 3 kb DNA fragment from the Streptomyces globisporus 1912 landomycin E (LaE) biosynthetic gene cluster (lnd) was completely sequenced. Three open reading frames were identified, lndGT4, lndZ4, and lndZ5, whose probable translation products resemble a glycosyltransferase, a reductase, and a hydroxylase, respectively. Studies of generated mutants from disruption and complementation experiments involving the lndGT4 gene allowed us to determine that LndGT4 controls the terminal L-rhodinose sugar attachment during LaE biosynthesis and that LndZ4/LndZ5 are responsible for the unique C11-hydroxylation of the landomycins. Generation of the novel landomycins F, G, and H in the course of these studies provided evidence for the flexibility of lnd glycosyltransferases toward their acceptor substrates and a basis for initial structure-activity relationships within the landomycin family of antibiotics.     

Modified AutoDock for accurate docking of protein kinase inhibitors

Protein kinases are an important class of enzymes controlling virtually all cellular signaling pathways. Consequently, selective inhibitors of protein kinases have attracted significant interest as potential new drugs for many diseases. Computational methods, including molecular docking, have increasingly been used in the inhibitor design process [1]. We have considered several docking packages in order to strengthen our kinase inhibitor work with computational capabilities. In our experience, AutoDock offered a reasonable combination of accuracy and speed, as opposed to methods that specialize either in fast database searches or detailed and computationally intensive calculations.However, AutoDock did not perform well in cases where extensive hydrophobic contacts were involved, such as docking of SB203580 to its target protein kinase p38. Another shortcoming was a hydrogen bonding energy function, which underestimated the attraction component and, thus, did not allow for sufficiently accurate modeling of the key hydrogen bonds in the kinase-inhibitor complexes.We have modified the parameter set used to model hydrogen bonds, which increased the accuracy of AutoDock and appeared to be generally applicable to many kinase-inhibitor pairs without customization. Binding to largely hydrophobic sites, such as the active site of p38, was significantly improved by introducing a correction factor selectively affecting only carbon and hydrogen energy grids, thus, providing an effective, although approximate, treatment of solvation.     

Chlorotrimethylsilane-promoted synthesis of 1,2,4-triazolopyrimidines from 3,5-diamino-1,2,4-triazoles and pentane-2,4-diones

An efficient synthesis of 2-amino-1-R-[1,2,4]triazolo[1,5-a]- pyrimidinium or 3-amino-2-R-[1,2,4]triazolo[4,3-a]pyrimidi- nium chloride derivatives by heterocyclization of 3,5-diamino- 1-R-1,2,4-triazoles (R = Alk or Ar) with pentane-2,4-diones was developed. The process is promoted by chlorotrimethyl- silane which plays the dual role of carbonyl-activating agent and water scavenger.     

Aggregation-Enhanced Emission and Thermally Activated Delayed Fluorescence of Derivatives of 9-Phenyl-9H-Carbazole: Effects of Methoxy and tert-Butyl Substituents

Derivatives of 9-phenyl-9H-carbazole were synthesized as efficient emitters exhibiting both thermally activated delayed fluorescence and aggregation-induced emission enhancement. Effects of methoxy and tert-butyl substituents at the different positions of carbazolyl groups on the properties of the emitters were studied. Depending on the substitutions, photoluminescence quantum yields (PLQY) of non-doped solid films of the compounds ranged from 17 % to 53 % which were much higher than those observed for the solutions in low-polarity solvent toluene. Compounds substituted at C-3 and C-6 positions of carbazole moiety by methoxy- and tert-butyl- groups showed the highest solid-state PLQY. Ionization potentials of the studied derivatives in solid-state were found to be in the short range of 5.75–5.89 eV. Well-balanced hole and electron mobilities were detected for tert-butyl-substituted compound. They exceeded 10⁻⁴ cm² (V×s)⁻¹ at electric fields higher than 3×10⁵ V cm⁻¹. Two compounds with the highest solid-state PLQYs showed higher efficiencies in non-doped organic light-emitting diodes than in the doped devices. Maximum external quantum efficiency of 7.2 % and brightness of 15000 cd m⁻² were observed for the best device.