title: Hybrid Mass Spectrometry Approaches to Determine How L-Histidine Feedback Regulates the Enzyzme MtATP-Phosphoribosyltransferase.
abstract: MtATP-phosphoribosyltransferase (MtATP-PRT) is an enzyme catalyzing the first step of the biosynthesis of L-histidine in Mycobacterium tuberculosis, and proposed to be regulated via an allosteric mechanism. Native mass spectrometry (MS) reveals MtATP-PRT to exist as a hexamer. Conformational changes induced by L-histidine binding and the influence of buffer pH are determined with ion mobility MS, hydrogen deuterium exchange (HDX) MS, and analytical ultracentrifugation. The experimental collision cross-section (DTCCSHe) decreases from 76.6 to 73.5 nm2upon ligand binding at pH 6.8, which correlates to the decrease in CCS calculated from crystal structures. No such changes in conformation were found at pH 9.0. Further detail on the regions that exhibit conformational change on L-histidine binding is obtained with HDX-MS experiments. On incubation with L-histidine, rapid changes are observed within domain III, and around the active site at longer times, indicating an allosteric effect.
authors.affiliation: Michael Barber Centre for Collaborative Mass Spectrometry, Manchester Institute of Biotechnology, School of Chemistry, 131 Princess Street, Manchester M1 7DN, UK; School of Chemistry, University of Edinburgh, David Brewster Road, Edinburgh EH9 3FJ, UK.
authors.forename: Kamila J
authors.initials: KJ
authors.lastname: Pacholarz
authors.affiliation: UCB Celltech, 216 Bath Road, Slough SL1 3WE, UK.
authors.forename: Rebecca J
authors.initials: RJ
authors.lastname: Burnley
authors.affiliation: The Biomolecular Analysis Facility, Faculty of Life Sciences, University of Manchester, Oxford Road, Manchester M13 9PT, UK.
authors.forename: Thomas A
authors.initials: TA
authors.lastname: Jowitt
authors.affiliation: School of Chemistry, University of Edinburgh, David Brewster Road, Edinburgh EH9 3FJ, UK.
authors.forename: Victoria
authors.initials: V
authors.lastname: Ordsmith
authors.affiliation: Mycobacterial Metabolism and Antibiotic Research Laboratory, The Francis Crick Institute, 1 Midland Road, London NW1 1AT, UK.
authors.forename: João Pedro
authors.initials: JP
authors.lastname: Pisco
authors.affiliation: School of Chemistry, University of Edinburgh, David Brewster Road, Edinburgh EH9 3FJ, UK.
authors.forename: Massimiliano
authors.initials: M
authors.lastname: Porrini
authors.affiliation: Mycobacterial Metabolism and Antibiotic Research Laboratory, The Francis Crick Institute, 1 Midland Road, London NW1 1AT, UK.
authors.forename: Gérald
authors.initials: G
authors.lastname: Larrouy-Maumus
authors.affiliation: UCB Celltech, 216 Bath Road, Slough SL1 3WE, UK.
authors.forename: Rachel A
authors.initials: RA
authors.lastname: Garlish
authors.affiliation: UCB Celltech, 216 Bath Road, Slough SL1 3WE, UK.
authors.forename: Richard J
authors.initials: RJ
authors.lastname: Taylor
authors.affiliation: Mycobacterial Metabolism and Antibiotic Research Laboratory, The Francis Crick Institute, 1 Midland Road, London NW1 1AT, UK. Electronic address: luiz.carvalho@crick.ac.uk.
authors.forename: Luiz Pedro Sório
authors.initials: LPS
authors.lastname: de Carvalho
authors.affiliation: Michael Barber Centre for Collaborative Mass Spectrometry, Manchester Institute of Biotechnology, School of Chemistry, 131 Princess Street, Manchester M1 7DN, UK. Electronic address: perdita.barran@manchester.ac.uk.
authors.forename: Perdita E
authors.initials: PE
authors.lastname: Barran
mesh_terms: D001276:ATP Phosphoribosyltransferase / Q000737:chemistry* / Q000378:metabolism; D000494:Allosteric Regulation; D000495:Allosteric Site; D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism; D025461:Feedback, Physiological; D006639:Histidine / Q000737:chemistry / Q000378:metabolism; D013058:Mass Spectrometry / Q000379:methods; D009169:Mycobacterium tuberculosis / Q000201:enzymology; D011485:Protein Binding
chemical_list: D001426:Bacterial Proteins; D006639:Histidine; D001276:ATP Phosphoribosyltransferase
keywords: ATP-phosphoribosyltransferase; allostery; hydrogen deuterium exchange; ion mobility; protein conformation; structural mass spectrometry; tuberculosis