title: Structural insights into the loss of catalytic competence in pectate lyase activity at low pH.
abstract: Pectate lyase, a family 1 polysaccharide lyase, catalyses cleavage of the α-1,4 linkage of the polysaccharide homogalacturonan via an anti β-elimination reaction. In the Michaelis complex two calcium ions bind between the C6 carboxylate of the d-galacturonate residue and enzyme aspartates at the active centre (+1 subsite), they withdraw electrons acidifying the C5 proton facilitating its abstraction by the catalytic arginine. Here we show that activity is lost at low pH because protonation of aspartates results in the loss of the two catalytic calcium-ions causing a profound failure to correctly organise the Michaelis complex.
authors.affiliation: Institut Laue Langevin, 71 Avenue des Martyrs, 38042 Grenoble Cedex 9, France; School of Biological and Chemical Sciences, Queen Mary University of London, Department of Chemistry & Biochemistry, Mile End Road, London E1 4NS, United Kingdom.
authors.forename: Salyha
authors.initials: S
authors.lastname: Ali
authors.affiliation: Department of Chemistry, University of Copenhagen, 2100 Copenhagen, Denmark.
authors.forename: Chresten R
authors.initials: CR
authors.lastname: Søndergaard
authors.affiliation: Institut Laue Langevin, 71 Avenue des Martyrs, 38042 Grenoble Cedex 9, France; EPSAM, Keele University, Staffordshire ST5 5BG, United Kingdom.
authors.forename: Susana
authors.initials: S
authors.lastname: Teixeira
authors.affiliation: School of Biological and Chemical Sciences, Queen Mary University of London, Department of Chemistry & Biochemistry, Mile End Road, London E1 4NS, United Kingdom. Electronic address: r.w.pickersgill@qmul.ac.uk.
authors.forename: Richard W
authors.initials: RW
authors.lastname: Pickersgill
mesh_terms: D001120:Arginine / Q000737:chemistry*; D001224:Aspartic Acid / Q000737:chemistry*; D001412:Bacillus subtilis / Q000201:enzymology*; D001426:Bacterial Proteins / Q000737:chemistry / Q000378:metabolism; D001665:Binding Sites; D002118:Calcium / Q000737:chemistry; D020134:Catalytic Domain; D006863:Hydrogen-Ion Concentration; D008958:Models, Molecular; D011133:Polysaccharide-Lyases / Q000737:chemistry* / Q000378:metabolism*; D017433:Protein Structure, Secondary; D013379:Substrate Specificity
chemical_list: D001426:Bacterial Proteins; D001224:Aspartic Acid; D001120:Arginine; D011133:Polysaccharide-Lyases; C023911:pectate lyase; D002118:Calcium
keywords: Catalysis; Loss of calcium-binding; Michaelis complex; Pectate lyase; pH dependence of activity