Graham R. Moran
Ph.D., University of New England, Australia
My research interests are primarily concerned with the activation of molecular oxygen by enzymes. Specifically, enzymes that catalyze oxygenations of aromatic compounds.
In recent years, the events leading to the activation of molecular oxygen by enzymes have become clearer. However, many of the subsequent catalytic events in which highly reactive oxo-intermediates are directed to react along a single coordinate to give a specific product remain to be elucidated. In the trapping and identification of these transients is a current frontier of enzymology.
Enzymes that react with molecular oxygen need some mediator to carry out the reduction/activation step. These mediators are either an organic prosthetic group capable of one electron chemistry, or a metal ion such as Fe(II), Fe(III), Mn(II) or Cu(II). We are currently concerned with the structure and catalysis of three oxygenase enzymes. Two of these use Fe(II) as a mediator and one uses flavin adenine dinucleotide.
An enzyme in the pathway for the catabolism of tyrosine, 4-hydroxyphenylpyruvate dioxygenase (HPPD) catalyze the incorporation of both atoms of molecular oxygen into an aromatic substrate bringing about a substantial rearrangment. The range of reactions include decarboxylation, hydroxylation, and subsituent migration in a single catalytic cycle.
HPPD has particularly diverse chemistry. The reaction catalysed is intriguing due to the considerable alteration that occurs in the conversion of its substrate, hydroxyphenylpyruvate (HPP) to its product HG. But also the inhibition of the enzyme in man eliminates the symptoms of three debilitating or lethal diseases. Inhibition of the enzyme in plants stops the production of plastoquinone and tocopherol and thus uncouples photosynthesis and kills the plant.
Hydroxymandelate synthase (HMS) is a mechanistic homolog of HPPD involved in the production of phenylglycine which is key component of non-protein peptide antibiotics such as vancomycin and chloroemycin. The differences in HPPD and HMS may reveal how reactive oxo-intermediates can be directed along specific paths.
Kynurenine-3-monooxygenase (KMO) is a flavoprotein hydroxylase that controls the levels of quinolinic acid, 3-hydroxykynurenine, xanthurenic acid and kynureninic acid. All of these molecules play key roles in the outcome of ischemic trauma in the brain. Our objective is to learn enough about the KMO reaction mechanism such that specific inhibitors can be devised that will allow modulation of the levels of these molecules for the treatment of stroke, epilepsy, and alziemers disease.
The methods we employ tend be based in kinetics. Our approaches measure individual rate constants for specific reaction steps. Presteady-state analysis using specialized rapid reaction techniques such as stopped-flow and rapid quench is being used to elucidate the chemical steps involved in catalysis or inhibition. These investigations are augmented by a series of steady-state methods and a variety of static spectroscopic measurements.
Adrienne R. Diebold, Michael L. Neidig, Graham R. Moran, Grit D. Straganz, Edward I. Solomon. (2010). "The 3His triad in Dke1: Comparisons to the Facial Triad." Biochemistry, 49, 6945-6952.
June Brownlee, Brian Heinz, Judith Bates, Graham R. Moran. (2010). "Product Analysis and Inhibition Studies of the Causative Asn->Ser Variant of 4-Hydroxyphenylpyruvate Dioxygenase Suggest a Simple Route to the Treatment of Hawkinsinuria." Biochemistry, 49, 7218-7226.
Panqing He, John A. Conrad and Graham R. Moran. (2010). "Rate limiting processes in the catalysis of Hydroxymandelate Synthase from Amycolatopsis orientalis." Biochemistry, 49, 1998-2007.
Panqing He and Graham R. Moran. (2009). "We Two Alone Will Sing: The Two-Substrate α-Keto Acid Dependent Oxygenases." Current Opinion in Chemical Biology, 13, 436-443.
Karen R. Crozier-Reabe, Robert S. Phillips and Graham R. Moran. (2008). "Kynurenine 3-Monooxygenase from Pseudomonas fluorescens: Substrate-Like Inhibitors Both Stimulate Flavin Reduction and Stabilize the Flavin-peroxo Intermediate yet Result in the Production of Hydrogen Peroxide." Biochemistry, 47, 12420–12433.
John A. Conrad and Graham R. Moran. (2008). "The Interaction of Hydroxymandelate Synthase with 4-Hydroxyphenylpyruvate Dioxygenase Inhibitors." Inorganica Chimica Acta, 361, 1197-1201.
June Brownlee, Panqing He, Graham R. Moran, and David H. T. Harrison (2008). "Two Roads Diverged: The Structure of Hydroxymandelate Synthase from Amycolatopsis orientalis in Complex with 4-Hydroxmandelate." Biochemistry, 47, 2002-2013.
Karen E. Keith, Lauren Killip, Panqing He, Graham R. Moran and Miguel A. Valvano. (2007). "Burkholderia cenocepacia C5424 Produces a Pigment with Antioxidant Properties Using a Homogentisate Intermediate." Journal of Bacteriology, 189, 9057-9065.
Vincent M. Purpero and Graham R. Moran. (2007). "The Diverse and Pervasive Chemistries of the α–Keto Acid Dependent Enzymes." Journal of Biological Inorganic Chemistry, 12, 587-601.
Karen R. Crozier and Graham R. Moran (2007). "Heterologous Expression and Purification of Kynurenine-3-monooxygenase from Pseudomonas fluorescens strain 17400." Protein Expression and Purification, 51, 324-333.