Dr. Ildiko M. Kovach
Emeritas, Bio-Organic Chemistry.
B. S., 1964, Szemmelweisz University of Medical Sciences,
Budapest, Hungary. Ph. D., 1974,
University of Kansas. Postdoctoral Fellow, 1974-1978, University of Kansas.
Tel: (202) 319-6550
Summary of Research Interests
The most fundamental idea pursued in our research program is the origin and evolutionary progress of catalytic acceleration by enzymes. The level of molecular recognition and the catalytic potential of active functional groups in small proteins and serine protease enzymes are probed. Specific targets are the cardiovascular enzymes and the cholinesterases, which are of great scientific and medical interest.
A central question addressed in our studies is the role of short-strong-hydrogen bonds (SSHBs) in catalysis and molecular recognition. A key target of this project is human ?-thrombin inhibited by a variety of small and large peptide inhibitors. The quest for understanding catalytic efficiency has in recent years included the role of general protein dynamics and their influence on remote-site interactions that are critical in the response of α-thrombin to large protein or peptide natural substrates and other effectors. Acetylcholinesterase as an evolutionarily highly developed catalyst has also been the subject of many investigations aimed at evaluating the importance of optimization of the step for leaving group departure in hydrolytic enzymes and the sensitivity of the catalytic machinery to alterations in the electronic and steric environments of substrates and inhibitors.
Our studies span the fields of physical organic chemistry (particularly isotope effects), biochemistry (particularly mechanistic enzymology) and, occasionally, organic synthesis. Experimental tools include stopped-flow kinetics, spectroscopic, fluorometric and electro-chemical methods. High-resolution 1H NMR, especially its applications to the characterization of deshielded SSHBs, is used in collaboration. Complementary structural information is obtained with molecular modeling of macromolecule-small molecule interactions by computer graphics combined with various energy minimizations and molecular dynamics calculations. Funding of these efforts have been provided in recent years by the NIH, American Heart Association and DOD.
I. M. Kovach, P. Kelley, C. Eddy, F. Jordan, and A. Baykal, Proton Bridging in the Interaction of Thrombin with Small Inhibitors. Biochemistry 48, 7296-7304 (2009).
D. Zhang and I. M. Kovach and J. P. Sheehy Locating the Rate-Determining Step for Three-Step Hydrolase-Catalyzed Reactions with DYNAFIT. Biochimica at Biophysica Acta: Proteins and Proteomics 1784, 827-833, (2008).
D. Zhang and I. M. Kovach Deuterium Solvent Isotope Effect and Proton Inventory Studies of Factor
Xa-Catalyzed Reactions. Biochemistry 45, 14175-14182, (2006).
D. Zhang and I. M. Kovach Full and Partial Deuterium Solvent Isotope Effect Studies of a-Thrombin-Catalyzed Reactions of Natural Substrates, J. Am. Chem. Soc. 127, 3760-3766 (2005).
E. J. Enyedy and I. M. Kovach Proton Inventory Studies of a-Thrombin-catalyzed Reactions of Substrates
with Selected P and P? Sites. J. Am. Chem. Soc. 126, 6017-6024 (2004).
I. M. Kovach Stereochemistry and Secondary Reactions in the Irreversible Inhibition of Serine Hydrolases
by Organophosphorus Compounds. J. Physical Organic Chemistry 17, 602-614 (2004).
A.S. Mildvan, M.A. Massiah, T.K. Harris, G.T. Marks, D.H. T. Harrison, C. Viragh, P. M. Reddy and
I. M. Kovach Short Strong Hydrogen Bonds on Enzymes: NMR and Mechanistic
Studies. J. Mol. Structure 163-175, (2002).
P. M. Reddy and I. M. Kovach Synthesis of chiral methyl 4-nitrophenyl alkylphosphonothioates:
BF3-Et2O-catalyzed alcoholysis of phosphonamidothioates. Tetrahedron Letters 43, 4063-4066 (2002).
P. M. Reddy, C. Viragh and I. M. Kovach Synthesis of Phenyl or 4-Nitrophenyl Methyl b-Ketophosphonate
Esters. Phosphorus Sulfur and Silicon 177, 1-4, (2002).
A.S. Mildvan, M.A. Massiah, T.K. Harris, G.T. Marks, D.H. T. Harrison, C. Viragh, P. M. Reddy
and I. M. Kovach Short Strong Hydrogen Bonds on Enzymes: NMR and Mechanistic Studies.
J. Mol. Structure (2002).
I. M. Kovach Inhibition of Serine Hydrolases by Organophosphorus Compounds. Biokemia XXV, 74-80
(Hungarian invited review) 2001.
I. J. Enyedy and I. M. Kovach, A. Bencsura, A Molecular Dynamics Study of Active-site Interactions with
Tetracoordinate Transients in Acetylcholinesterase and its Mutants. Biochemical J. 353, 645-653 (2001).
M. A. Massiah, C. Viragh, P. M. Reddy, I. M. Kovach, J. Johnson, T. L. Rosenberry and A. S. Mildvan,
Short, Strong Hydrogen Bonds at the Active Site of Human Acetylcholinesterase: 1 H-NMR Studies.
Biochemistry 40, 5682-5690 (2001).
C. Viragh, T. K. Harris, P. M. Reddy, M. A. Massiah, A. S. Mildvan and I. M. Kovach, NMR Evidence
for a Short, Strong Hydrogen Bond in the Mechanism of a Cholinesterase. Biochemistry 39,
E. Enyedy and I. M. Kovach, Reversible Modulation of Human Factor Xa Activity by Phosphonate
Ester: Media Effects. Bioorg. Med. Chem. 8, 549-556 (2000).
C. Viragh, I. M. Kovach and Lewis Pannell, Small Molecular Products of Dealkylation in Soman-inhibited
Electric Eel Acetylcholinesterase. Biochemistry 38, 9557 (1999).
I. M. Kovach, Ligand and Active-Site Dependent P-O Versus C-O Bond Cleavage in Organophosphorus Adducts of
Serine Hydrolases. Phosphorus, Sulfur and Silicon, 144-146, 537-540 (1999).
A. Saxena, C. Viragh, D. S. Frazier, I. M. Kovach, D. M. Maxwell, O. Lockridge and B. P. Doctor, The pH Dependence of Dealkylation in Soman-inhibited Cholinesterases and Their Mutants: Further Evidence for a Push-Pull Mechanism. Biochemistry 37, 15086-15096 (1998).
I. J. Enyedy, I. M. Kovach and B. R. Brooks, Alternate Pathways for Acetic Acid and Acetate Ion Release from Acetylcholinesterase: A Molecular Dynamics Study. J. Am. Chem. Soc. 120, 8043-8050 (1998).
I. M. Kovach and E. Enyedy , Active-site-dependent Elimination of 4-Nitrophenol from 4-Nitrophenyl
Alkylphosphonyl Serine Protease Adducts. J. Am. Chem. Soc. 120, 258-263 (1998).
I. M. Kovach, R. Akhmetshin, I. J. Enyedy and C. Viragh A Selfconsistent Mechanism for Dealkylation in
Soman-inhibited Acetylcholinesterase. Biochemical J. 324, 995-996 (1997).
C. Viragh, R. Akhmetshin, I. M. Kovach, and C. Broomfield, Unique Push-Pull Mechanism of Ageing in
Soman-inhibited Acetylcholinesterase. Biochemistry 36, 8243-8252 (1997).
E. Enyedy and I. M. Kovach, Modulation of the Activity of Human α-Thrombin with Phosphonate Ester
Inhibitors. Bioorg. Med. Chem. 5, 1531-1541, (1997).
A. Bencsura, I. Enyedy and I. M. Kovach, Probing the Active Site of Acetylcholinesterase by Molecular
Dynamics of its Phosphonate Ester Adducts. J. Am. Chem. Soc. 118, 8531-8541 (1996).
I. Enyedy, A. Bencsura and I. M. Kovach, Interactions in Tetravalent and Pentavalent Phosphonate Esters
of Ser at the Active Site of Serine Enzymes. Phosphorus, Sulfur, and Silicon 109-110, 249-252 (1996).
Q. Zhao and I. M. Kovach, Reversible Modification of Tissue-type Plasminogen Activator by
Methylphosphonate Esters . Bioorg. Med. Chem 4, 523-529 (1996).
A. Bencsura, I. Enyedy and I. M. Kovach, Origins and Diversity of the Aging Reaction in Phosphonate
Adducts of Serine Hydrolase Enzymes: What Characteristics of the Active Site Do They Probe?
Biochemistry 34, 8989-8999 (1995).
I. M. Kovach, N. Qian and A. Bencsura, Efficient Product Clearance through Exit Channels in Substrate
Hydrolysis by Acetylcholinesterase. FEBS Lett. 349, 60-64 (1994).
A. Saxena, N. Qian, I. M. Kovach, A. P. Kozikowski, Y. P. Pang, D. C. Vellom, Z. Radic, D.
Quinn, P. Taylor, and B. P. Doctor, Identification of Amino Acid Residues Involved in the Binding
of Huperzine-A to Cholinesterases. Protein Sci., 1770-1778 (1994).
Q. Zhao, I.M. Kovach, A. Bencsura and A. Papathanassiu, Enantioselective and Reversible Inhibition
of Trypsin and α-Chymotrypsin by Phosphonate Esters. Biochemistry 33, 8128 (1994).
N. Qian and I. M. Kovach, Key Active Site Residues in the Inhibition of Acetylcholinesterases by Soman.
FEBS Lett. 336, 263 (1993).
I. M. Kovach, Q. Zhao, M. Keane and R. Reyes, Rate-Determining Carbonyl Hydration in the Intramolecular
Hydrolysis of Phenacyl Phosphonate Esters: Isotopic Probes and Activation Parameters.
J. Am. Chem. Soc.. 115, 10471 (1993).
I. M. Kovach, A.J. Bennet, J.A. Bibbs, and Q. Zhao, Nucleophilic and Protolytic Catalysis of
Phosphonate Ester Hydrolysis: Isotope Effects and Activation Parameters.
J. Am. Chem. Soc. 115, 5138 (1993).
I. M. Kovach Solution Dynamics of Phosphonate Ester Hydrolysis. Phosphorus,
Sulfur and Silicon 75, 131 (1993).
I. M. Kovach, L. McKay and D. Vander Velde, Diastereomeric Phosphonate Ester Adducts of
Chymotrypsin: 31P-NMR Measurements. Chirality 5, 143 (1993).
I. M. Kovach and L. McKay Reversible Modulation of Serine Protease Activity by Phosphonate Esters.
Bioorg. Med. Chem. Lett. 2, 1735 (1992).
I. M. Kovach and D. Huhta Comparative Study of the Charge Distribution in Tetravalent
Carbonyl Transients and Organophosphorus Adducts of Trypsin. J. Mol. Struct.
(Theochem 79) 233, 335 (1991).
I. M. Kovach, Competitive Irreversible Inhibition of Enzymes in the Presence of a Substrate: Scope and
Limitations. J. Enzyme Inhibit. 4, 201 (1991).
I. M. Kovach, D. Huhta and S. Baptist Active Site Interactions in Hydrated Trypsin-Organophosphate
Adducts: A YETI Molecular Mechanics Study. J. Mol. Struct. (Theochem 72) 226, 99 (1991).
I. M. Kovach, Son Do and R.L. Schowen β-Secondary Deuterium Isotope Effect and Solvent Isotope
Effects in Catalysis by Subtilisin BPN'. J. Phys. Org. Chem. 3, 260 (1990).
I. M. Kovach and A.J. Bennet Comparative Study of Nucleophilic and Enzymic Reactions of 2-Propyl
Methylphosphonate Derivatives. Phosphorus, Sulfur and Silicon 51/52, 51 (1990).
Recent invited presentations
“Short Strong Hydrogen Bonds at the Active Site of Cholinesterases” I. M. Kovach, Gordon Research
Conference on Isotopes in Biological and Chemical Sciences, Ventura, CA, February 17-22, 2002.
“Short Strong Hydrogen Bonds at the Active Site of Cholinesterases: 1H-NMR Studies” I. M. Kovach, C. Viragh, P. M. Reddy, M. A. Massiah, A. S. Mildvan, J. Johnson and T. L. Rosenberry, VIIth International Meeting on Cholinesterases, Pucon, Chile, November 8-12, 2002.
“Protonic Participation in the Rate-determining Step for the Hydrolysis of Fluorogenic Substrates Catalyzed by Hemostatic and Thrombolytic Enzymes.” I. M. Kovach, E. J. Enyedy and R. A. Akhmetshin, International Isotope Effects Conference an EUCHEM Conference, Uppsala, Sweden, June 22-27, 2003.
"Protonic Participation in Hemostatic and Thrombolytic Enzyme-catalyzed Reactions." I. M. Kovach,
E.J. Enyedy and R.A. Akhmetshin, XVth International Conference on Horizons in Hydrogen Bond
Research, Freie Universitat Berlin, Germany, September 16-21, 2003.
“Full and Partial Deuterium Solvent Isotope Effect Studies of a-Thrombin-Catalyzed Reactions of Natural Substrates” D. Zhang and I. M. Kovach, Isotopes 2005 Bath: An international conference, The University of Bath, UK, June 27- July 1, 2005.
"Short Strong Hydrogen Bonds in Support of the Inhibition of Human a-Thrombin." I. M. Kovach,
Gordon Research Conference on Isotopes in Biological and Chemical Sciences, Galveston, TX, February 14-19, 2010.
Fogarty Fellowship, Hungarian National Academy of Sciences, Institute of Enzymology ,
Senior Faculty Research Associateship in the Navy-ASEE program. Laboratory for the
Structure of Matter, Naval Research Laboratory, Washington , DC
NSF Career Advancement Award for Women
American Heart Association, Nation's Capital Affiliate 1992 Heart Ball Research Award
NIH/NIA Individual National Research Service Award, Laboratory of Structural Biology,
DCRT, NIH, Bethesda, MD
NRC Senior Research Associateship, Division of Biochemistry, Walter Reed Army Institute
of Research, Washington, DC.
Editor and Member of the Editorial Board of the Biochemical Journal.
Senior Fulbright lecturing award, 2003-2004 host institute Department of Biochemistry as
Eotvos Lorand University.