Allostery is defined as the intramolecular control that involves the effect of one ligand on the binding or catalysis of another with no direct interaction between them. Meanwhile cooperativity is defined as the mutual assistance of interactions of the ligand with a macromolecule at different equivalent binding sites. Then, signal propagation between binding sites is dependent on the tertiary and quaternary structure of the protein.
Allostery and cooperativity in proteins have been associated in most regulated proteins. Both behaviours are well explained, at phenomenological level of detail, by the general model of Monod, Wyman, and Changeux (MWC). In this model, regulated proteins are oligomers, that show only one active site per monomer, and display thermodynamic coupling between both equivalent (homotropic) and non-equivalent (heterotropic) sites by a reversible transition between two discrete states. In spite of the robustness of MWC model, some new mechanisms have been found due to present day computational methods, combined with structural, thermodynamic, and kinetic studies, which make possible a different approach to provide a deeper understanding of the requirements for allostery.
GlcN6P deaminase (EC 3.5.99.6, NagB) in the enzyme which catalyzes the isomerization-deamination of Glucosamine-6-phosphate (GlcN6P) releasing Fructose-6-phosphate (Fru6P) and ammonium ion (NH4+). This enzyme plays a crucial role in amino-sugar metabolism and, in some species, is an essential metabolic regulatory step since it displays positive cooperativity and allosteric activation by N-Acetylglucosamine-6-phosphate (GlcNAc6P). In E. coli, NagB is a homohexamer, with a Rossmann-like fold, usually referred as NagBI. Nevertheless, NagB from the bacterium Shewanella oneidensis and the archaeon Thermococcus kodakaraensis present an entirely different fold, based on the sugar isomerase (SIS) domain. This fold is found in several proteins that have a role in phosphosugar isomerization, like glucosamine-fructose-6- phosphate aminotransferase (GlmS).
In S. oneidensis, the analogous of NagBI was designated NagBII and it also displays positive cooperativity and allosteric activation by GlcNAc6P, representing an unusual case of homoplasy since the convergence is not only in its catalytic function, but also in its allosteric regulation and cooperativity. Convergence on functional and regulatory mechanisms in similar selective contexts from two different structures strongly suggests adaptation, and it highlights the importance of allosteric enzymes in control mechanisms of the cell.
Herein we present the kinetic properties of NagBII from S. denitrificans (SdNagBII) and the crystallographic structures of the free enzyme as well as the enzyme complexed with 2-amino-2-deoxy-glucitol-6-phosphate (GlcNol6P) alone or with the allosteric activator GlcNAc6P. Fitting kinetic data of this enzyme to Hill equation revealed that the intensity of substrate cooperativity is too high for a dimer, revealing the existence of a non-previously described mechanism of cooperativity. Differential binding of these molecules clearly explain the origin of the intense cooperativity found for this enzyme through a non-previously described mechanism of cooperativity.