BBB seminar: Albert Mildvan
Structures and mechanisms of nucleoside-diphosphate-X (NUDIX) hydrolases
Department of Biological Chemistry, Johns Hopkins University School of Medicine, Baltimore, MD, USA
Nudix hydrolases catalyze the hydrolysis of nucleoside diphosphates linked to other moieties, X, and contain the sequence motif or Nudix box, GX5EX7REUXEEXGU. The mechanisms of 8-oxo-dGTPase (MutT), and of GDP-mannose hydrolase will be discussed in detail, as typical examples.
Nudix enzymes are found in all organisms where they play protective, regulatory, and signaling roles in metabolism. The mechanisms of action of Nudix hydrolases are highly diverse in the position on the substrate at which bond cleavage occurs, and in the number of required divalent cations. While most proceed by associative nucleophilic substitutions by water at specific internal phosphorus atoms of a diphosphate or polyphosphate chain, members of the GDP-mannose hydrolase sub-family catalyze dissociative nucleophilic substitutions, by water, at carbon. The site of substitution is determined by the positions of the general base and the entering water. The rate accelerations or catalytic powers of Nudix hydrolases range from 109- to 1012-fold. The reactions are accelerated 103- to 105-fold by general base catalysis by a glutamate residue within, or beyond the Nudix box, or by a histidine beyond the Nudix box. Lewis acid catalysis, which contributes ≥103- to 105-fold to the rate acceleration, is provided by one, two, or three essential divalent cations. One divalent cation is coordinated by two or three conserved residues of the Nudix box, the initial glycine and one or two glutamate residues, together with a remote glutamate or glutamine ligand from beyond the Nudix box. Additional catalysis (10- to 103-fold) is provided by the cationic side chains of lysine and arginine residues and by H-bond donation by tyrosine residues, to orient the general base, or to promote the departure of the leaving group. The overall rate accelerations can be explained by both independent and cooperative effects of these catalytic components.
Host: Aurora Martínez, Department of Biomedicine
|Albert Mildvan, M.D., Professor of Biological Chemistry and Chemistry, focuses his research interests on the mechanisms of enzyme action. In order to understand the structural basis for the high catalytic power of enzymes, he and his group study the structures and mechanisms of enzyme-catalyzed hydrolytic reactions of nucleotides, as well as tautomeric isomerases and cholinesterases. Multidimensional heteronuclear NMR, electron spin resonance, chemical and genetic modification of enzymes, and kinetic isotope effects are used to elucidate the solution structures of enzymes, the roles of metals, the presence of strong hydrogen bonds, and the conformations, interactions, arrangement, and exchange rates of enzyme-bound substrates.|