A neurosteroid analogue photolabeling reagent labels the colchicine-binding site on tubulin: a mass spectrometric analysis

Previous studies have shown that the neurosteroid analogue, 6-Azi-pregnanolone (6-AziP), photolabels voltage-dependent anion channels and proteins of approximately 55 kDa in rat brain membranes. The present study used two-dimensional electrophoresis and nanoelectrospray ionization ion-trap mass spectrometry (nano-ESI-MS) to identify the 55 kDa proteins (isoelectric point 4.8) as isoforms of β-tubulin. This identification was confirmed by immunoblot and immunoprecipitation of photolabeled protein with anti-β-tubulin antibody and by the demonstration that 6-AziP photolabels purified bovine brain tubulin in a concentration-dependent pattern. To identify the photolabeling sites, purified bovine brain tubulin was photolabeled with 6-AziP, digested with trypsin, and analyzed by matrix-assisted laser desorption/ionization MS (MALDI). A 6-AziP adduct of TAVCDIPPR(m/z?= 1287.77), a β-tubulin specific peptide, was detected by MALDI. High-resolution liquid chromatography-MS/MS analysis identified that 6-AziP was covalently bound to cysteine 354 (Cys-354), previously identified as a colchicine-binding site. 6-AziP photolabeling was inhibited by 2-methoxyestradiol, an endogenous derivative of estradiol thought to bind to the colchicine site. Structural modeling predicted that neurosteroids could dock in this colchicine site at the interface between α- and β-tubulin with the photolabeling group of 6-AziP positioned proximate to Cys-354.     

Powder Pattern Hyperfine Spectroscopy of Shallow- Donor Muonium Centres

The hyperfine spectroscopy of muonium in II–VI semiconductors is reviewed, suggesting that whereas hydrogen is a deep-level defect in ZnS, ZnSe and ZnTe, it constitutes a shallow donor in ZnO, CdS, CdSe and CdTe. Shallow and deep states coexist in CdTe. Using new data for ZnO, it is shown that the principal values of the muonium hyperfine tensor may be obtained with equal facility from measurements in longitudinal or in transverse magnetic field, and from samples that are polycrystalline powders or single crystals. Spin density on the central muon in the shallow states correlates with the electron binding energy or donor depth. This revised version was published online in September 2006 with corrections to the Cover Date.     

Magnetism and candidate muon-probe sites in RBa2Cu3O y

Key results of zero-field (ZF) and transverse-field (TF) muon-spin-relaxation (μSR) experiments on superconducting and insulating RBa₂Cu₃O _y (R123 _y , with R=Eu, Gd, Pr and Pr/Y:y=6, 7) are examined. The chemical behavior of the positive muon probe is addressed, and muon-oxygen bonding is shown to occur in all these cuprates. To explain magnetic fields at muon-probe sites in Pr _x Y_1−x Ba₂Cu₃O _y (0<=x<0.5,y=7 andx=0,y=6) samples, improvements on the reported magnetic structures from neutron diffraction are necessary. Cu magnetism in Pr123y (y=6,7) is observed belowT _N1, which is near RT. The magnetism seen belowT _N2 can be interpreted assuming an additional ordering in the Cutt-O chain layers. Alternatively, Pr ordering is also considered as the cause of the second phase transition. Considering the specific muon-probe location, a more detailed interpretation can be provided for the μSR parameters, measured in the normal and mixed states of these unconventional superconductors.     

Diffusion, trapping, and relaxation of Mu+ and Mu- in heavily‐doped GaAs

The diamagnetic muonium states in heavily doped GaAs are investigated with a combination of transverse‐field and longitudinal‐field μSR techniques. In metallic n‐type GaAs, formation of Mu^- occurs because of the high Fermi energy. This analog of the hydride ion (H^-) is located in a T_Ga interstice where it is essentially immobile up to about 500 K. At higher temperatures, Mu_T acts as an electron–hole recombination center. In p‐type GaAs, Mu⁺ traps at two different sites, one at low temperatures and a second at higher temperatures after detrapping from the first. This revised version was published online in August 2006 with corrections to the Cover Date.     

Development of a μLCR facility at LAMPF

It has long been recognized thatLCR could profitably be done with the high intensity surface beam at LAMPF [1]. A spectrometer has been built that is matched to the LAMPF beam characteristics. The polarization information is obtained from a downstream array of counters while side counters, containing no polarization signal, monitor the ⁺ beam. Degraders select higher energy e⁺, thereby reducing rates and required counter segmentation while maintaining information content. We apply a ramped longitudinal field in addition to the static one to average over instabilities in the ⁺ beam. This field scan allows direct interpretation of data and does not require a prior estimate of the resonance structure of a sample. Flux coils monitor the applied ramp field and eddy-current induced fields. High average rate (2×10^{7 +}/s). good stability, and the versatile field scan permitted useful data to be collected from Cu, Al(Cu), Al, Si(Al), and polycrystalline Si targets.     

Muonium dynamics in doped Si above room temperature studied by longitudinal field — μSR

We report longitudinal fieldSR 1/T ₁ measurements in Si from room temperature to 850 K. The data in pure Si and SiB (p-type) can be explained in a two-state model where muonium cycles between its positive and neutral charge states. Within this model, the average muon-electron hyperfine parameter in the neutral state is consistent with muonium at the tetrahedral interstitial site, indicating that at the highest temperatures measured, neutral muonium spends a significant amount of time away from the bond centered site, the calculated potential minimum. Although this is also true for SiP (n-type) at high temperatures, the data in the region between 300–450 K indicates that at least one other state is involved in the dynamics.This work is partially supported by the Natural Sciences and Engineering Research Council of Canada, the Welch Foundation (C-1048 [TLE], D-1053 [RLL] and the U.S. National Science Foundation (DMR-8917639 [TLE,BH]).     

Die Struktur von Cyclosporin C

The Structure of Cyclosporin C The structure of cyclosporin C ( 2 ), a minor antifungal metabolite from Trichoderma polysporum (Link ex Pers.) RIFAI has been elucidated. Hydrolytic cleavage and spectroscopic evidence show that cyclosporin C is a neutral oligopeptide of 11 amino acids linked together in a 33-membered ring. Cyclosporin C ( 2 ) differs from the main metabolite cyclosporin A ( 1 ) [2] [4] only by containing L-threonine in the place of L-α-aminobutyric acid as has been shown by the conversion of 2 into 1 . ¹³C-NMR. spectra and study of molecular models suggest that cyclosporin C ( 2 ) has the same molecular conformation as 1 , which is best described as a twisted β-pleated sheet held together in a conformationally stable form by intramolecular hydrogen bonding.     

A study of structural transformation rates in NiTiF6·6D2O

The progression rate versus temperature for the supercooled structural transformation in NiTiF₆·6D₂O is compared to that of NiTiF₆·6H₂O. Energy parameters are obtained from a fit to a nucleation and growth model. Deuteration lowers the critical temperature by ≈ 14 K, and a second transition seen in the hydrated salt is not unambiguously observed.