Targeted mass spectrometric strategy for global mapping of ubiquitination on proteins

Post-translational modifications of proteins including phosphorylation, glycosylation, acetylation and ubiquitination facilitate the regulation of many cellular processes and intracellular signaling events. Ubiquitination plays a key role in the functional regulation and degradation of many classes of proteins, and the study of ubiquitination and poly-ubiquitination has emerged as one of the most active areas in proteomic research. A variety of mass spectrometric methods have been described for the identification of ubiquitination sites, the study of poly-ubiquitin topology and the identification of ubiquitin substrates. The most popular workflow for both ubiquitination site mapping and poly-ubiquitination chain topology characterization is to take advantage of the Gly-Gly signature on the substrate's lysine residue observed after tryptic digestion. Although a number of protocols have been described for the mapping of ubiquitination sites, one major challenge is that ubiquitination is typically heterogeneous, and several lysine residues may be ubiquitinated within a protein. When multiple ubiquitination sites are present, multiple analyses are often required to cover all of the potential modification sites which in turn can necessitate the usage of larger quantities of material. In addition, the level of ubiquitination on endogenous and recombinant proteins may be of low intensity, adding further analytical challenges in the identification of this modification. The use of the multiple reaction monitoring (MRM)-initiated detection and sequencing workflow (MIDAS) for the identification of phosphorylation sites has previously been described. Here, we explore the use of an MRM workflow for ubiquitination site mapping on the substrate protein, receptor interacting protein (RIP).     

Swift heavy ion-induced effects in Ce-doped nickel ferrite nanoparticles

Swift heavy ions of various energies are being used for material modifications. The induced modifications depend on the kind of defects produced during interaction of ions with the target material. In the present work, irradiation of 200 MeV Ag beam-induced effects in NiFe₂O₄ and NiCe_0.04Fe_1.96O₄ nanoparticles are studied at two different fluences, 2×10¹² and 1×10¹³ ions/cm². Nanoparticles of nickel ferrite and Ce-doped nickel ferrite were prepared by chemical route. X-ray diffraction pattern shows peaks corresponding to pure spinel structure in both the systems, NiFe₂O₄ and NiCe_0.04Fe_1.96O₄. The pristine as well as irradiated nanoparticles were characterized by high-resolution transmission electron microscopy (HRTEM), Raman spectroscopy, electron paramagnetic resonance spectroscopy (EPR) and vibrating sample magnetometer (VSM). Raman spectra show bands corresponding to spinel structure. After irradiation, the position of the bands does not change significantly for both samples. The widths corresponding to the same band in both the systems show opposite trend with fluence. VSM results show that after irradiation, the magnetization decreases from 40 to 32 A m²/kg for NiFe₂O₄ and from 39 to 31 A m²/kg for NiCe_0.04Fe_1.96O₄. EPR results show that after doping with Ce as well as irradiation, the EPR line width is reduced, making samples important for applications.     

Enantioresolution of Some β-Blockers and a β2-Agonist Using Ligand Exchange TLC

JPC – Journal of Planar Chromatography – Modern TLC - Cu(II) complexes of l-threonine, l-serine, and l-tartaric acid were prepared and used as ligand exchange reagents for enantiomeric...     

Amino acids as chiral selectors in enantioresolution by liquid chromatography

Amino acids are unique in terms of their structural features and multidimensional uses. With their simple structures and the ready availability of both enantiomers, amino acids not only serve as a chiral pool for synthesis but also provide an inexpensive pool for resolution studies. There has been no attempt to review the application of amino acids as chiral selectors for chromatographic enantioresolution of pharmaceuticals and other compounds. The present paper deals with application of l-amino acids and complexes of l-amino acids with a metal ion, particularly Cu(II), as an impregnating reagent in thin-layer chromatography or as a chiral ligand exchange reagent or a chiral mobile phase additive in both thin-layer chromatography and high-performance liquid chromatography. Enantiomeric resolution of β-blockers, nonsteroidal anti-inflammatories, amino acids (and their derivatives) and certain other compounds is discussed.     

HPLC enantioresolution of (R,S)-baclofen using three newly synthesized dichloro-s-triazine reagents having amines and five others having amino acids as chiral auxiliaries

Enantioresolution of (R,S)-baclofen was accomplished using a newly synthesized set of three chiral derivatizing reagents (CDRs) having amines [(S)-(-)-α,4-dimethylbenzylamine, (-)-cis-myrtanylamine and (R)-(-)-1-cyclohexylethylamine] as chiral auxiliaries in cyanuric chloride and another set of five CDRs having amino acids (L-Leu, D-Phg, L-Val, L-Met and L-Ala) as chiral auxiliaries. These eight CDRs were used for synthesis of diastereomers of (R,S)-baclofen under microwave irradiation. The diastereomers were separated on a reversed-phase C(18) column using mixtures of methanol with aqueous trifluoroacetic acid with UV detection at 230 nm. Chromatographic data obtained for the two sets of diastereomers were compared among themselves and among the two groups. The method was validated for limit of detection, linearity, accuracy and precision.     

Orbital Valence,Urey-Bradley and Approximate General Valence Force Fields and Vibrational Mean Amplitudes of Some Spherical Top M(CH3)4 Type Molecular Model (M=C,Si, Ge,Sn and Pb)

The intramolecular force fields of tetramethylcarbon, tetramethylsilane, tetramethylgermanium, tetramethyltin and tetramethyllead have been examined using OVFF, UBFF and GVFf potential functions under the approximation of ‘point-mass’ for methyl group. Approximate force fields for the (2x2)F₂ species of each molecule have been examined using L₁₂=O, L₂₁=O and P.E.D., methods. Transferability of OVFF into GVFF has also been observed. An analysis of the Coriolis coupling constants has been presented in the light of the force fields of the molecules. Vibrational mean amplitudes for bonded and non-bonded distances have been reported and analysed. The Bastiansen-Morino shrinkage effedt has been evaluated. The results have been discussed in the light of the relative behaviour of the molecules.     

Quantitative Analysis by Derivative Electronic Spectroscopy

Abstract

Absorption and emission spectroscopy in the ultraviolet and visible regions, apart from being the earliest physical techniques of analysis, have great utility in solving a variety of structural and analytical problems [l-41. However, in many cases the quantitative determination of individual components in a mixture by UV-VIS spectroscopy becomes very difficult owing to the spectral similarities and overlapping of weak absorption bands of one component by strong absorption bands of other components. It was recently recognized that derivative spectrophotometry, introduced about three decades ago [5-81 and originally proposed by Rutherford [9], can be a very useful analytical tool for characterizing an analyte band that is overlapped by other absorption bands with different halfwidths. On differentiation, the intensity of the absorption band with a small halfwidth increases more than that of a band with a large halfwidth [10]. The central idea behind the development of the science of derivatized spectra rests upon the measurement of the first or the higher derivatives of the absorbance A or the luminescence intensity I with respect to the wavelength λ, e.g., dI/dλ, dA/dλ, d²I/dλ², etc. These derivatives, when plotted or scanned against wavelength, yield an array of derivative spectra.     

Analysis on the laser beam pointing instability induced fringe shift and contrast dilution from different interferometers used for writing fiber Bragg grating

This paper presents an analysis on the effect of laser beam pointing instability on the fringe shift and hence the contrast dilution of superposed fringes from two beam interferometers. The interferometers analyzed are those commonly used in writing fiber Bragg grating (FBG) such as phase mask, bi-prism and phase mask based Talbot/Holographic type. The beam pointing instability is incorporated as slight change in the angle between two interfering beams. The relative immunity of different interferometers to laser beam pointing is discussed vs location of FBG writing plane from the beam splitter. The effect of the beam pointing was minimum in proximity FBG writing by phase mask. The effect, in terms of fringes contrast dilution, was worst in case of large length interferometers e.g. phase mask based-Talbot interferometer. For intermediate length prism interferometers, the effect was moderate. For a given length interferometer, the fringe shift was directly proportional to beam pointing angle and inversely proportional to fringe separation. Theoretical analysis is verified experimentally by studying the fringe instability of interference pattern formed by a bi-prism of angle 2^o with the copper vapour laser (CVL, λ = 510 nm) beams of different beam pointing instabilities.     

New physics, the cosmic ray spectrum knee, and pp cross section measurements

We explore the possibility that a new-physics interaction can provide an explanation for the knee just above 10⁶ GeV in the cosmic ray spectrum. We model the new-physics modifications to the total proton–proton cross section with an incoherent term that allows for missing energy above the scale of new physics. We add the constraint that the new physics must also be consistent with published pp cross section measurements, using cosmic ray observations, an order of magnitude and more above the knee. We find that the rise in cross section required at energies above the knee is radical. The increase in cross section suggests that it may be more appropriate to treat the scattering process in the black disc limit at such high energies. In this case there may be no clean separation between the standard model and new-physics contributions to the total cross section. We model the missing energy in this limit and find a good fit to the Tibet III cosmic ray flux data. We comment on testing the new-physics proposal for the cosmic ray knee at the Large Hadron Collider.     

Synthesis,Antimicrobial Evaluation,and Docking Studies of Substituted Acetylphenoxymethyl‐triazolyl‐N‐phenylacetamides

A series of molecules containing acetylphenoxymethyl, triazole, and N‐phenylacetamide moieties were synthesized via the click chemistry approach. All the synthesized compounds were screened for their antimicrobial activities in vitro. The synthesized compounds 8a , 8b , 8m , and 8n showed better activities. We further performed exploratory docking studies to gain some insight regarding the molecular mechanism of antibacterial action of these compounds that could guide further structure‐activity relationship (SAR) studies. We examined the interaction of the most active compound with DNA gyrase (pdb id:1KZN). Based on antimicrobial and docking studies, the compounds 8a , 8b , 8m , and 8n were identified as potential antimicrobial agents.