research papers 10 Rudin Äo-Pin Äera et al.  Structural flexibility in glucosamine-6-phosphate deaminase Acta Cryst. (2002). D58, 10±20 Acta Crystallographica Section D Biological Crystallography ISSN 0907-4449 Structural flexibility, an essential component of the allosteric activation in Escherichia coli glucosamine-6-phosphate deaminase E. Rudin Äo-Pin Äera,* S. Morales- Arrieta,² S. P. Rojas-Trejo and E. Horjales Departamento de Reconocimiento Molecular y Bioestructura, Instituto de Biotecnologõ Âa, Universidad Nacional Auto  noma de Mexico, PO Box 510-3, Cuernavaca, MOR 62271, Mexico ² Present address: Centro de Investigaciones Biolo  gicas del Noroeste, Mar Bermejo 195, La Paz, BCS 23090, Mexico. Correspondence e-mail: rudino@ibt.unam.mx # 2002 International Union of Crystallography Printed in Denmark ± all rights reserved A new crystallographic structure of the free active-site R conformer of the allosteric enzyme glucosamine-6-phosphate deaminase from Escherichia coli, coupled with previously reported structures of the T and R conformers, generates a detailed description of the heterotropic allosteric transition in which structural ¯exibility plays a central role. The T conformer's external zone [Horjales et al. (1999), Structure, 7, 527±536] presents higher B values than in the R conformers. The ligand-free enzyme (T conformer) undergoes an allosteric transition to the free active-site R conformer upon binding of the allosteric activator. This structure shows three alternate conformations of the mobile section of the active-site lid (residues 163±182), in comparison to the high B values for the unique conformation of the T conformer. One of these alternate R conformations corresponds to the active-site lid found when the substrate is bound. The disorder associated with the three alternate conformations can be related to the biological regulation of the K m of the enzyme for the reaction, which is metabolically required to maintain adequate concentrations of the activator, which holds the enzyme in its R state. Seven alternate conformations for the active-site lid and three for the C-terminus were re®ned for the T structure using isotropic B factors. Some of these conformers approach that of the R conformer in geometry. Furthermore, the direction of the atomic vibrations obtained with an- isotropic B re®nement supports the hypothesis of an oscillating rather than a tense T state. The concerted character of the allosteric transition is also analysed in view of the apparent dynamics of the conformers. Received 5 March 2001 Accepted 8 October 2001 PDB References: R form glucosamine-6-phosphate deaminase complex with fructose 6-phosphate at 2.15 A Ê , 1fqo; R form glucos- amine-6-phosphate deami- nase complex with N-acetyl- glucosamine-6-phosphate at 2.10 A Ê , 1frz; R form glucos- amine-6-phosphate deami- nase complex with N-acetyl- glucosamine-6 phosphate at 1.73 A Ê , 1fs5; T form glucos- amine-6-phosphate deami- nase at 2.2 A Ê , 1fs6; T form glucosamine-6-phosphate deaminase at 1.9 A Ê , 1fsf. 1. Introduction Allosteric regulation involves communication between distant ligand-binding sites on biological macromolecules, which is central to many cellular regulatory mechanisms (Perutz, 1990). It encompasses two kinds of transitions: (i) homotropic, in which binding of a molecule to one subunit modulates the binding of the same type of molecule to the other subunits, and (ii) heterotropic, in which binding of a ligand in its speci®c site triggers a conformational change that modulates the binding properties of a second type of molecule in the binding site (Goodsell & Olson, 2000). In allosteric enzymes, this control process involves two structurally different states which differ in quaternary structure: the T form (`tense form'), in which the af®nity for the substrate is zero or low, and the R form (`relaxed form') with an optimal af®nity for substrate and/or allosteric activator. The metabolic regulation achieved by allosteric enzymes results from their ability to shift the