Dissecting the mechanism by which glycosyltransferases calalyse mannosyl transfer
Lead Research Organisation:
University of Oxford
Department Name: Oxford Chemistry
Abstract
The linking of sugars to a variety of different proteins has an important influence on the function of cells. The biological catalysts or enzymes that speed up the reactions in which sugars are linked to other molecules are known as glycosyltransferases. Although these enzymes and are industrially and biologically important, they have not been extensively studied because they are difficult to produce in large amounts. In this project we will take advantage of our ability to produce significant quantities of glycosyltransferases that catalyse the transfer of the sugar mannose onto other molecules. The three dimensional structure of these enzymes will be determined and the information will be used to synthesise inhibitors of these biological catalysts and to use molecular engineering techniques to manipulate their biological properties.
Technical Summary
Although mannose-containing polymers are widespread in nature, there is a paucity of structural and mechanistic information on the enzymes that catalyze mannosyltransfer. Recent studies by our three groups [Flint et al. (2005) Nat. Struct. Mol. Biol. 12, 608-14] have begun to unravel the structural basis for the catalytic activity and plasticity of substrate recognition of the retaining GDP-Man transferase, mannosylglycerate synthase, which lays a foundation upon which to dissect mannosyl transfer. This application will build upon our studies on mannosylglycerate synthase and a significant additional body of preliminary data, of both retaining and inverting mannosyltransferases (including one of the key enzymes of glycobiology, dolichyl-phosphate -D-mannose synthase) to dissect the mechanism of action and specificity of mannosyltransferases. This will underpin the engineering of these enzymes to increase their utility as biosynthetic tools, underpin novel therapeutic strategies that target glycan decoration and, and, through the modulation of key enzymes that catalyse mannosyltransfer, provide profound insights into cellular function To date, there are no known selective inhibitors of any retaining glycosyltransferases with sufficient potency to allow modulation of the function of these enzymes in vivo. This void has hampered not only the understanding of the role of mannose decoration in biology, but also the exploitation of mannosyl transfer in drug design. The key goals that will be addressed in this project are: (a) Determine how structure dictates specificity and the mechanism of catalysis of mannosyltransferases (b) Exploitation of such information in the design of new enzyme inhibitors which reflect both structural and mechanistic features (c) Interrogation of the evolution of the mechanisms of mannosyl transfer, and its exploitation in the development of novel biocatalysts.
Publications
A Celik
(2007)
Direct Biocatalytic Conversion of Carbon Dioxide: Towards a Formic Acid Economy
in in preparation
A Celik
(2007)
Creating new Tools for a Formic Acid Economy: Cloning & Structure of Novel Dehydrogenases
in in preparation
Aljabali A. A. A.
(2014)
Multimeric microparticles of iron oxide (mMPIO) for dual modality molecular imaging with CT & MRI
in EUROPEAN HEART JOURNAL
Allman SA
(2009)
Potent fluoro-oligosaccharide probes of adhesion in Toxoplasmosis.
in Chembiochem : a European journal of chemical biology
Baboo S
(2014)
Most human proteins made in both nucleus and cytoplasm turn over within minutes.
in PloS one
Backus KM
(2011)
Uptake of unnatural trehalose analogs as a reporter for Mycobacterium tuberculosis.
in Nature chemical biology
Backus KM
(2014)
The three Mycobacterium tuberculosis antigen 85 isoforms have unique substrates and activities determined by non-active site regions.
in The Journal of biological chemistry
Bagi Zsolt
(2013)
Microparticles bearing RGD peptide promote integrin avß3-dependent platelet adhesion in isolated, pressurized cerebral artery
in FASEB JOURNAL
Barry C
(2013)
'Naked' and Hydrated Conformers of the Conserved Core Pentasaccharide of N-linked Glycoproteins and Its Building Blocks
in Journal of the American Chemical Society
Barry CS
(2011)
ESI-MS assay of M. tuberculosis cell wall antigen 85 enzymes permits substrate profiling and design of a mechanism-based inhibitor.
in Journal of the American Chemical Society
Bernardes G
(2007)
A trisulfide-linked glycoprotein
in Chemical Communications
Bernardes GJ
(2008)
Facile conversion of cysteine and alkyl cysteines to dehydroalanine on protein surfaces: versatile and switchable access to functionalized proteins.
in Journal of the American Chemical Society
Bernardes GJ
(2011)
Site-selective traceless Staudinger ligation for glycoprotein synthesis reveals scope and limitations.
in Chembiochem : a European journal of chemical biology
Bernardes GJ
(2008)
From disulfide- to thioether-linked glycoproteins.
in Angewandte Chemie (International ed. in English)
Bernardes Goncalo J. L.
(2012)
Chemical site-selective protein modification: Development of a traceless vascular targeting ADC for cancer therapy
in ABSTRACTS OF PAPERS OF THE AMERICAN CHEMICAL SOCIETY
Bernardes Goncalo J. L.
(2012)
Building well-defined glycoproteins
in ABSTRACTS OF PAPERS OF THE AMERICAN CHEMICAL SOCIETY
Bolam DN
(2007)
The crystal structure of two macrolide glycosyltransferases provides a blueprint for host cell antibiotic immunity.
in Proceedings of the National Academy of Sciences of the United States of America
Boutureira O
(2009)
Accessible sugars as asymmetric olefin epoxidation organocatalysts: glucosaminide ketones in the synthesis of terminal epoxides.
in Organic & biomolecular chemistry
Boutureira O
(2012)
Selenenylsulfide-linked homogeneous glycopeptides and glycoproteins: synthesis of human "hepatic Se metabolite A".
in Angewandte Chemie (International ed. in English)
Boutureira O
(2010)
Fluoroglycoproteins: ready chemical site-selective incorporation of fluorosugars into proteins.
in Chemical communications (Cambridge, England)
Boutureira O
(2011)
Direct radiolabelling of proteins at cysteine using [18F]-fluorosugars.
in Chemical communications (Cambridge, England)
Boutureira Omar
(2012)
Chemical site-selective radiolabelling of proteins using fluorosugars
in ABSTRACTS OF PAPERS OF THE AMERICAN CHEMICAL SOCIETY
Campo VL
(2007)
Chemical and chemoenzymatic synthesis of glycosyl-amino acids and glycopeptides related to Trypanosoma cruzi mucins.
in Organic & biomolecular chemistry
Chalker J
(2011)
Methods for converting cysteine to dehydroalanine on peptides and proteins
in Chemical Science
Chalker JM
(2012)
Conversion of cysteine into dehydroalanine enables access to synthetic histones bearing diverse post-translational modifications.
in Angewandte Chemie (International ed. in English)
Description | We developed a detailed understanding of the mechanisms of glycosyltransferases based on a chemical approach. These enzymes are widespread in biology but their mechanism for bond-formation has been poorly understood. By building and using suitable chemical probes, we were able to provide the first experimental evidence for a wholly unexpected mechanism that is rare in traditional chemistry (the SNi). In contrast to the chemistry found in the test tube, nature seems very capable of using this unusual reaction. This has had implications for how we think about these important enzymes and how we can design drugs that block them to treat certain diseases. |
Exploitation Route | Understanding how these enzymes work informs the corresponding biology and also allows the design of drugs for a class of enzymes that has been one of the most difficult to target to date. |
Sectors | Agriculture, Food and Drink,Chemicals,Education,Environment,Healthcare,Pharmaceuticals and Medical Biotechnology |
URL | http://users.ox.ac.uk/~dplb0149/index.html |
Description | BGD and group members have appeared on the radio, television, (BBD, Channel 5), science festivals around the world (Cheltenham, Kent, Edinburgh, Times Lit., Sydney) describing this work. We have given talks in schools to inspire the next generation. The work contributed strongly to the expansion of biocatalysts available for use in synthesis and license options on technology agreed with biotech e.g. Prozomix. |
First Year Of Impact | 2008 |
Sector | Chemicals,Healthcare,Pharmaceuticals and Medical Biotechnology |
Impact Types | Societal,Economic |
Company Name | Glycoform Ltd |
Description | drug delivery and glycoprotein specialist; biopharmaceuticals |
Impact | Employed >20 people over 10 years and provided a model for how synthetic protein drugs might be constructed and used. The technology for this company has now been used by major US companies. |
Website | http://isis-innovation.com/news/glycoform-ltd-improve-drug-delivery/ |