Acyl and phosphoryl group transfer reactions are essential parts of the biochemistry of living systems. Little is known about the influences of leaving group and nucleophile, and the effect of catalytic groups, on the mechanisms and transition states of these reactions. This work will address these questions using a variety of isotope effect studies. Preliminary work supports a change from a stepwise mechanism with a tetrahedral intermediate to a concerted mechanism for acyl transfer reactions of esters with good leaving groups. The specific goals of the work in this proposal are to: 1. Obtain detailed descriptions of the transition states for acyl and phosphoryl transfer reactions in solution and in enzymatic reactions. Comparisons of these will shed light on the effect of the enzymatic catalytic groups on transition state structure. 2. Analyze differences in mechanism and in transition state structure that arise from changes in the leaving group and in the nucleophile (changes in basicity or nucleophilicity, and in identity of the attaching atom, for example, oxygen versus sulfur nucleophiles) in solution and enzymatic reactions. 3. Delineate the boundaries of the limiting concerted vs. stepwise mechanisms as determined by properties of nucleophile and/or leaving group in the solution and in enzymatic reactions. 4. Determine whether activated substrates undergo enzymatic group transfer reactions by the same mechanisms as unactivated ones, or if the mechanisms differ, as they do in solution.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
First Independent Research Support & Transition (FIRST) Awards (R29)
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Physical Biochemistry Study Section (PB)
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Utah State University
Schools of Arts and Sciences
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Hengge, Alvan C (2015) Kinetic isotope effects in the characterization of catalysis by protein tyrosine phosphatases. Biochim Biophys Acta 1854:1768-75
Kuznetsov, Vyacheslav I; Hengge, Alvan C (2013) New functional aspects of the atypical protein tyrosine phosphatase VHZ. Biochemistry 52:8012-25
Whittier, Sean K; Hengge, Alvan C; Loria, J Patrick (2013) Conformational motions regulate phosphoryl transfer in related protein tyrosine phosphatases. Science 341:899-903
Brandão, Tiago A S; Johnson, Sean J; Hengge, Alvan C (2012) The molecular details of WPD-loop movement differ in the protein-tyrosine phosphatases YopH and PTP1B. Arch Biochem Biophys 525:53-9
Kuznetsov, Vyacheslav I; Alexandrova, Anastassia N; Hengge, Alvan C (2012) Metavanadate at the active site of the phosphatase VHZ. J Am Chem Soc 134:14298-301
Kuznetsov, Vyacheslav I; Hengge, Alvan C; Johnson, Sean J (2012) New aspects of the phosphatase VHZ revealed by a high-resolution structure with vanadate and substrate screening. Biochemistry 51:9869-79
Brandão, Tiago A S; Hengge, Alvan C; Johnson, Sean J (2010) Insights into the reaction of protein-tyrosine phosphatase 1B: crystal structures for transition state analogs of both catalytic steps. J Biol Chem 285:15874-83
Miti?, Natasa; Hadler, Kieran S; Gahan, Lawrence R et al. (2010) The divalent metal ion in the active site of uteroferrin modulates substrate binding and catalysis. J Am Chem Soc 132:7049-54
Feng, Guoqiang; Tanifum, Eric A; Adams, Harry et al. (2009) Mechanism and transition state structure of aryl methylphosphonate esters doubly coordinated to a dinuclear cobalt(III) center. J Am Chem Soc 131:12771-9
Brandão, Tiago A S; Robinson, Howard; Johnson, Sean J et al. (2009) Impaired acid catalysis by mutation of a protein loop hinge residue in a YopH mutant revealed by crystal structures. J Am Chem Soc 131:778-86

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