Studies on the amino acid sequence of cytochrome c

The purpose of this work was to obtain information about the sequence of amino acid residues in the peptide chain of cytochrome c, and at the same time to study methods for obtaining ouch information. Cytochrome c Was isolated from horse heart d purified by chromatography on ion exchange resin (Amberlite IRC 50) and on sephadex. Attempts to identify the N-terminal amino acid residue yielded negative results, and it was concluded that the protein may not have a free terminal amino group. This was subsequently confirmed by Kreil and Tuppy who showed that the N-terminal group is acetylglycine. To investigate the internal sequence of the protein, cytochrome c was hydrolysed with trypsin to yield a mixture of peptides. Chromatography on carboxymethyl cellulose, brought about partial resolution of this mixture, and the application of high voltage paper electrophoresis permitted the isolation of some pure homogeneous peptides from the material thus obtained. These methods, however, seemed inadequate for the exhaustive analysis of the peptide mixture. An effort was made, therefore, to reduce the severity of the problem of peptide fractionation, by reducing the complexity of the mixture to be separated. This was achieved by acylating the lysine amino groups of cytochrome c, and thus restricting the action of trypsin to linkages involving the carbonyl groups of the two arginine residues in the protein. Methoxy-carboxyl chloride was firs tried as an acylating agent, but proved unsuitable. Ethyl thiotrifluoroacetate was satisfactory and, at pH 9, this ester reacted with all the amino groups of the protein. It was found also that the substituent trifluoroacetyl groups could subsequently be removed by means of 0.5 M ammonia, without affecting the peptide structure. The action of trypsin on the trifluoroacetyl protein yielded a mixture of peptides which was rather more complex the had been anticipated. On the assumption that this was due to the action of traces of chymotrypsin in the trypsin preparation the time of exposure to the protease was reduced, and it was eventually possible by means of chromatography on sephadex to isolate a small number of large peptides. The amino acid compositions of these peptides, and the nature of their N-terminal residues were determined. Also, one of them was hydrolysed with chymotrypsin, and the resulting mixture of small peptides fractionated on a column of ZeoKarb 225 x 2. The peptides thus separated were analysed and their sequence in the original peptide determined. The results of the analysis carried out during the course of this work were internally consistent, and in general found agreement with the recently published no acid sequence for cytochrome c

Isomorphic controllers and Dynamic Tuning: invariant fingering over a tuning continuum

The tuning invariance is where the relationship among the intervals of a given scale remain the same over a range of tunings but requires that the frequency differences are glossed over to expose the similarities. Tuning invariance can be a musically useful property by enabling dynamic tuning which is the real-time changes to the tuning of all sounded notes as a tuning variable changes along a smooth continuum. The mathematical and perceptual abstractions that are the prerequisite of this dynamic tuning are greatly discussed. Other topics being discussed include the identification of the note layouts that are tuning invariant, the meaning of the "same" across a range of tunings for a given interval and the definition of "range of tunings" for a given temperament

Measuring the effects of fractionated radiation therapy in a 3D prostate cancer model system using SERS nanosensors.

Multicellular tumour spheroids (MTS) are three-dimensional cell cultures that possess their own microenvironments and provide a more meaningful model of tumour biology than monolayer cultures. As a result, MTS are becoming increasingly used as tumor models when measuring the efficiency of therapies. Monitoring the viability of live MTS is complicated by their 3D nature and conventional approaches such as fluorescence often require fixation and sectioning. In this paper we detail the use of Surface Enhanced Raman Spectroscopy (SERS) to measure the viability of MTS grown from prostate cancer (PC3) cells. Our results show that we can monitor loss of viability by measuring pH and redox potential in MTS and furthermore we demonstrate that SERS can be used to measure the effects of fractionation of a dose of radiotherapy in a way that has potential to inform treatment planning.EaStCHEM, NHS Lothian, Jamie King Cancer Research FundThis is the final version of the article. It first appeared from the Royal Society of Chemistry via http://dx.doi.org/10.1039/C6AN01032

Targeted SERS nanosensors measure physicochemical gradients and free energy changes in live 3D tumor spheroids.

Use of multicellular tumor spheroids (MTS) to investigate therapies has gained impetus because they have potential to mimic factors including zonation, hypoxia and drug-resistance. However, analysis remains difficult and often destroys 3D integrity. Here we report an optical technique using targeted nanosensors that allows in situ 3D mapping of redox potential gradients whilst retaining MTS morphology and function. The magnitude of the redox potential gradient can be quantified as a free energy difference (ΔG) and used as a measurement of MTS viability. We found that by delivering different doses of radiotherapy to MTS we could correlate loss of ΔG with increasing therapeutic dose. In addition, we found that resistance to drug therapy was indicated by an increase in ΔG. This robust and reproducible technique allows interrogation of an in vitro tumor-model's bioenergetic response to therapy, indicating its potential as a tool for therapy development.Leverhulme Trust (Grant ID: RPG-2012-680), Jamie King Cancer Research FundThis is the final version of the article. It first appeared from the Royal Society of Chemistry via http://dx.doi.org/10.1039/C6NR06031

Convolutional neural networks for automated targeted analysis of gas chromatography-mass spectrometry data

Through their breath, humans exhale hundreds of volatile organic compounds (VOCs) that can reveal pathologies, including many types of cancer at early stages. Gas chromatography–mass spectrometry (GC-MS) is an analytical method used to separate and detect compounds in the mixture contained in breath samples. The identification of VOCs is based on the recognition of their specific ion patterns in GC-MS data, which requires labour-intensive and time-consuming preprocessing and analysis by domain experts. This paper explores the original idea of applying supervised machine learning, and in particular convolutional neural networks (CNNs), to learn ion patterns directly from raw GC-MS data. The method adapts to machine specific characteristics, and once trained, can quickly analyse breath samples bypassing the time-consuming preprocessing phase. The CNN classification performance is compared to those of shallow neural networks and support vector machines. All considered machine learning tools achieved high accuracy in experiments with clinical data from participants. In particular, the CNN-based approach detected the lowest number of false positives. The results indicate that the proposed method is a promising tool to improve accuracy, specificity, and in particular speed in the detection of VOCs of interest in large-scale data analysis

Towards an Intelligent Tutor for Mathematical Proofs

Computer-supported learning is an increasingly important form of study since it allows for independent learning and individualized instruction. In this paper, we discuss a novel approach to developing an intelligent tutoring system for teaching textbook-style mathematical proofs. We characterize the particularities of the domain and discuss common ITS design models. Our approach is motivated by phenomena found in a corpus of tutorial dialogs that were collected in a Wizard-of-Oz experiment. We show how an intelligent tutor for textbook-style mathematical proofs can be built on top of an adapted assertion-level proof assistant by reusing representations and proof search strategies originally developed for automated and interactive theorem proving. The resulting prototype was successfully evaluated on a corpus of tutorial dialogs and yields good results.Comment: In Proceedings THedu'11, arXiv:1202.453

The Atacama Cosmology Telescope: Cosmological parameters from three seasons of data

We present constraints on cosmological and astrophysical parameters from high-resolution microwave background maps at 148 GHz and 218 GHz made by the Atacama Cosmology Telescope (ACT) in three seasons of observations from 2008 to 2010. A model of primary cosmological and secondary foreground parameters is fit to the map power spectra and lensing deflection power spectrum, including contributions from both the thermal Sunyaev-Zeldovich (tSZ) effect and the kinematic Sunyaev-Zeldovich (kSZ) effect, Poisson and correlated anisotropy from unresolved infrared sources, radio sources, and the correlation between the tSZ effect and infrared sources. The power ell^2 C_ell/2pi of the thermal SZ power spectrum at 148 GHz is measured to be 3.4 +\- 1.4 muK^2 at ell=3000, while the corresponding amplitude of the kinematic SZ power spectrum has a 95% confidence level upper limit of 8.6 muK^2. Combining ACT power spectra with the WMAP 7-year temperature and polarization power spectra, we find excellent consistency with the LCDM model. We constrain the number of effective relativistic degrees of freedom in the early universe to be Neff=2.79 +\- 0.56, in agreement with the canonical value of Neff=3.046 for three massless neutrinos. We constrain the sum of the neutrino masses to be Sigma m_nu < 0.39 eV at 95% confidence when combining ACT and WMAP 7-year data with BAO and Hubble constant measurements. We constrain the amount of primordial helium to be Yp = 0.225 +\- 0.034, and measure no variation in the fine structure constant alpha since recombination, with alpha/alpha0 = 1.004 +/- 0.005. We also find no evidence for any running of the scalar spectral index, dns/dlnk = -0.004 +\- 0.012.Comment: 26 pages, 22 figures. This paper is a companion to Das et al. (2013) and Dunkley et al. (2013). Matches published JCAP versio

Asymmetric Wolbachia Segregation during Early Brugia malayi Embryogenesis Determines Its Distribution in Adult Host Tissues

Wolbachia are required for filarial nematode survival and fertility and contribute to the immune responses associated with human filarial diseases. Here we developed whole-mount immunofluorescence techniques to characterize Wolbachia somatic and germline transmission patterns and tissue distribution in Brugia malayi, a nematode responsible for lymphatic filariasis. In the initial embryonic divisions, Wolbachia segregate asymmetrically such that they occupy only a small subset of cells in the developing embryo, facilitating their concentration in the adult hypodermal chords and female germline. Wolbachia are not found in male reproductive tissues and the absence of Wolbachia from embryonic germline precursors in half of the embryos indicates Wolbachia loss from the male germline may occur in early embryogenesis. Wolbachia rely on fusion of hypodermal cells to populate adult chords. Finally, we detect Wolbachia in the secretory canal lumen suggesting living worms may release bacteria and/or their products into their host