Timothy Robert Rudd

Publications (32) View all

• Article: Antithrombin Stabilisation by Sulfated Carbohydrate s Correlates with Anticoagulant Activity

  Marcelo A. Lima, Ashley J. Hughes, Noemi Veraldi, Timothy R. Rudd, Rohanah Hussain, Adriana S. Brito, Suely F. Chavante, Ivarne I. Tersariol, Giuliano Siligardi, Helena B. Nader, Edwin A. Yates
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  ABSTRACT: Thermal stabilisation of native antithrombin-III (AT), determined using differential scanning fluorimetry, correlated with the anticoagulant activity of heparin and heparin-related saccharides, while similar conformational changes were induced in native AT by a variety of active and inactive heparin-related sulfated carbohydrates, measured in solution using synchrotron radiation circular dichroism, and their anticoagulant activities. Measurement of native AT stabilisation provides a convenient assay for prospective anticoagulants and represents an additional parameter by which to compare biosimilar heparins.
  Medicinal Chemistry Communications. 04/2013;
• Article: Analysis of the FGFR Signalling Network with Heparin as Co-Receptor: Evidence for the expansion of the core FGFR signaling network.

  Ruoyan Xu, Timothy R Rudd, Ashley J Hughes, Giuliano Siligardi, David G Fernig, Edwin A Yates
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  ABSTRACT: Evolution of the fibroblast growth factor (FGF) - FGF receptor (FGFR) signaling system follows closely that of multicellular organisms. The abilities of 9 FGFs (FGFs 1-9; examples of FGF sub-families -1, -4, -7, -8 and -9) and 7 FGF receptors or isoforms (FGFR1b, 1c, 2b, 2c, 3b, 3c and 4) to support signaling in the presence of heparin, a proxy for the cellular heparan sulfate co-receptor, were assembled into a network. A connection between two FGFRs was defined as their mutual ability to signal with a particular FGF. The network contained a core of 4 receptors (FGFR1c, 2c, 3c and 4) with complete connectivity and high redundancy. Analysis of the wider network indicated that neither FGF-3 nor FGF-7 was well-connected to this core of 4 receptors and, that divergence of a precursor of FGF sub-groups 1, 4 and 9 from FGF subgroup 8 may have allowed expansion from a 3 member FGFR core signaling system to the 4 member core network. This increases 4-fold the number of possible signaling combinations. Synchrotron Radiation Circular dichroism spectra of the FGFs with heparin revealed no overall common structural change, suggesting distinct heparin binding sites throughout the FGFs. The approach provides a potential method of identifying agents capable of influencing particular FGF/FGFR combinations, or areas of the signaling network for experimental, or therapeutic purposes. © 2013 The Authors Journal compilation © 2013 FEBS.
  FEBS Journal 02/2013; · 3.79 Impact Factor
• Article: An unusual antithrombin-binding heparin octasaccharide with an additional 3-O-sulfated glucosamine in the active pentasaccharide sequence.

  Marco Guerrini, Stefano Elli Sr, Pierre Mourier, Timothy R Rudd, Davide Gaudesi, Benito Casu, Christian Boudier, Giangiacomo Torri, Christian Viskov
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  ABSTRACT: The 3-O-sulfation of N-sulfated-glucosamine is the last event in the biosynthesis of heparin/heparan sulfate, giving rise to the antithrombin-binding pentasaccharide sequence AGA*IA, which is largely associated with the antithrombotic activity of these molecules. The aim of this study was the structural and biochemical characterisation of a previously unreported AGA*IA*-containing octasaccharide isolated from the very low molecular weight heparin semuloparin, in which both glucosamine residues of the pentasaccharide moiety located at the non-reducing end bear 3-O-sulfate groups. Two-dimensional and STD NMR experiments clearly confirmed its structure and identified its ligand epitope binding to antithrombin. The molecular conformation of the octasaccharide/antithrombin complex has been determined by NMR experiments and docking/energy minimisation. The presence of the second 3-O-sulfated glucosamine in the octasaccharide induced more than one order of magnitude increase in affinity to antithrombin compared to the pentasaccharide AGA*IA.
  Biochemical Journal 10/2012; · 4.90 Impact Factor
• Article: A zinc complex of heparan sulfate destabilises lysozyme and alters its conformation.

  Ashley J Hughes, Rohanah Hussain, Cesare Cosentino, Marco Guerrini, Giuliano Siligardi, Edwin A Yates, Timothy R Rudd
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  ABSTRACT: The naturally occurring anionic cell surface polysaccharide heparan sulfate is involved in key biological activities and is implicated in amyloid formation. Following addition of Zn-heparan sulfate, hen lysozyme, a model amyloid forming protein, resembled β-rich amyloid by far UV circular dichroism (increased β-sheet: +25%), with a significantly reduced melting temperature (from 68 to 58°C) by fluorescence shift assay. Secondary structure stability of the Zn-heparan sulfate complex with lysozyme was also distinct from that with heparan sulfate, under stronger denaturation conditions using synchrotron radiation circular dichroism. Changing the cation associated with heparan sulfate is sufficient to alter the conformation and stability of complexes formed between heparan sulfate and lysozyme, substantially reducing the stability of the protein. Complexes of heparan sulfate and cations, such as Zn, which are abundant in the brain, may provide alternative folding routes for proteins.
  Biochemical and Biophysical Research Communications 08/2012; 425(4):794-9. · 2.48 Impact Factor
• Article: How to find a needle (or anything else) in a haystack: two-dimensional correlation spectroscopy-filtering with iterative random sampling applied to pharmaceutical heparin.

  Timothy R Rudd, Eleonora Macchi, Cristina Gardini, Laura Muzi, Marco Guerrini, Edwin A Yates, Giangiacomo Torri
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  ABSTRACT: Risks of contamination of the major clinical anticoagulant heparin can arise from deliberate adulteration with unnatural or natural polysaccharides, including heparin from other animal sources, other natural products, or artifacts of manufacture, and these can escape detection by conventional means. Currently, there is no generally applicable, objective test recommended by regulators that can detect these in pharmaceutical heparin, and this continues to leave heparin exposed to contamination risks. Two-dimensional correlation spectroscopic-filtering with iterative random sampling (2D-COS-firs) is reported. It employs a difference covariance matrix with iterative random sampling, and is capable of revealing contamination in pharmaceutical heparin to a high level of sensitivity irrespective of the nature of those features. The technique is suitable to any situation in which a comparison of a single entity to a family of heterogeneous entities, particularly natural products and biosimilars, needs to be made, and will find application in pharmaceutical monitoring, manufacturing quality control, materials science, biotechnology, and metabolomic investigations.
  Analytical Chemistry 07/2012; 84(15):6841-7. · 5.86 Impact Factor