.

Department of Pathology, Microbiology, and Immunology

 

 

  Ingrid M. Verhamme, Ph.D.

 

 

Research Assistant Professor

Dept of Pathology, Microbiology and Immunology


 

  Contact Info
 


  Office Location:

T-2309

Phone: 615-343-6563  

E-mail: ingrid.verhamme@vanderbilt.edu

 

 Campus Email address:

 Pathology, Microbiology and Immunology-3rd Fl

 C-3321 MCN (2561)

 

 US Mailing address:

 Vanderbilt University School of Medicine

 Pathology, Microbiology and Immunology-3rd Fl

 C-3321 MCN

 Nashville, TN 37232-2561

 

 

 

Education

 

 

Ph.D., State Univ of Gent, Belgium
M.S. (Pharmacist), State Univ Gent
B.S., State Univ of Gent

 

 

Research Keywords

 

 

equilibrium binding, rapid and steady-state kinetics

 

 

Research Description

 

 

Our mechanism-based studies involve inactivation of the central clotting protease, thrombin, by the serine proteinase inhibitors (SERPINs) heparin cofactor II (HCII), antithrombin (AT) and plasminogen activator inhibitor-1 (PAI-1). These irreversible inactivation reactions are selectively accelerated by glycosaminoglycans (GAGs) such as heparin, dermatan sulfate, heparan sulfate, and in the case of PAI-1, also by vitronectin. Regulation of these reactions is important in controlling thrombosis and reocclusion.
Thrombin localized on fibrin, and on the cell surface GAGs reacts mainly with HCII and platelet PAI-1 at the injury site, in GAG-accelerated mechanisms different from AT. Subendothelial dermatan sulfate selectively enhances covalent T-HCII formation. Only HCII, in the presence of dermatan sulfate, inhibits fibrin-bound thrombin, by an unknown mechanism, and HCII has been identified as an important inhibitor of arterial thrombosis.
Thrombin possesses two Arg- and Lys-rich exosites, I and II. Exosite I is thought to bind HCII and PAI-1 directly, whereas exosite II - heparin interactions may modulate HCII and PAI-1 turnover as thrombin substrates. These mechanism steps are absent in the thrombin - AT reaction. PAI-1 stabilizes arterial thrombi, but paradoxically, may also promote embolization after atherosclerotic plaque rupture. Selective PAI-1 inhibitory vs. substrate activity toward localized thrombin may be a regulatory mechanism for modulating thrombin activity in arterial clots.

Our lab studies the mechanisms important in thrombin localization, inhibitory and substrate serpin recognition, and the mechanisms by which GAGs act as molecular switches for inhibiting or promoting serpin-mediated inactivation of fibrin-bound thrombin. In collaborative studies, we recently expanded our interests to factor XIa and XIIa interactions with AT and PAI-1, and to serpin-independent anticoagulant and profibrinolytic effects of GAGs, at the level of inhibition of prothrombinase activity, and enhancement of plasminogen activation by tPA and uPA.

Rapid and steady-state kinetics, and equilibrium binding with native and mutant proteins are used to characterize enzyme inactivation, serpin substrate pathway modulation, preferred pathways of productive complex formation, and regulation by fibrinogen and fibrin. These techniques allow quantitation of protein-protein and protein-GAG affinities, and conformational and chemical reaction steps. The combined approaches are aimed at quantitating the HCII and PAI-1 mechanisms of thrombin interaction in the presence of GAGs, and the effect of fibrin on these mechanisms. The goal of these studies is to understand differential HCII and PAI-1 regulation of clot-localized, thrombin-dependent processes in arterial thrombosis and reocclusion.

 

 

Publications

 

 

Verhamme, IM. Fluorescent reporters of thrombin, heparin cofactor II, and heparin binding in a ternary complex. Anal Biochem, 421(2), 489-98, 2012

Geng, Y, Verhamme, IM, Messer, A, Sun, MF, Smith, SB, Bajaj, SP, Gailani, D. A sequential mechanism for exosite-mediated factor IX activation by factor XIa. J Biol Chem, 287(45), 38200-9, 2012

Matafonov, A, Sarilla, S, Sun, MF, Sheehan, JP, Serebrov, V, Verhamme, IM, Gailani, D. Activation of factor XI by products of prothrombin activation. Blood, 118(2), 437-45, 2011

Sarilla, S, Habib, SY, Kravtsov, DV, Matafonov, A, Gailani, D, Verhamme, IM. Sucrose octasulfate selectively accelerates thrombin inactivation by heparin cofactor II. J Biol Chem, 285(11), 8278-89, 2010

Sarilla, S, Habib, SY, Tollefsen, DM, Friedman, DB, Arnett, DR, Verhamme, IM. Glycosaminoglycan-binding properties and kinetic characterization of human heparin cofactor II expressed in Escherichia coli. Anal Biochem, 406(2), 166-75, 2010

Smith, SB, Verhamme, IM, Sun, MF, Bock, PE, Gailani, D. Characterization of Novel Forms of Coagulation Factor XIa: independence of factor XIa subunits in factor IX activation. J Biol Chem, 283(11), 6696-705, 2008

Verhamme, IM, Bock, PE. Rapid-reaction kinetic characterization of the pathway of streptokinase-plasmin catalytic complex formation. J Biol Chem, 283(38), 26137-47, 2008

Bock, PE, Panizzi, P, Verhamme, IM. Exosites in the substrate specificity of blood coagulation reactions. J Thromb Haemost, 5 Suppl 1, 81-94, 2007

Panizzi, P, Boxrud, PD, Verhamme, IM, Bock, PE. Binding of the COOH-terminal lysine residue of streptokinase to plasmin(ogen) kringles enhances formation of the streptokinase.plasmin(ogen) catalytic complexes. J Biol Chem, 281(37), 26774-8, 2006

Bean, RR, Verhamme, IM, Bock, PE. Role of the streptokinase alpha-domain in the interactions of streptokinase with plasminogen and plasmin. J Biol Chem, 280(9), 7504-10, 2005

Ogawa, T, Verhamme, IM, Sun, MF, Bock, PE, Gailani, D. Exosite-mediated substrate recognition of factor IX by factor XIa. The factor XIa heavy chain is required for initial recognition of factor IX. J Biol Chem, 280(25), 23523-30, 2005

Boxrud, PD, Verhamme, IM, Bock, PE. Resolution of conformational activation in the kinetic mechanism of plasminogen activation by streptokinase. J Biol Chem, 279(35), 36633-41, 2004

Verhamme, IM, Bock, PE, Jackson, CM. The preferred pathway of glycosaminoglycan-accelerated inactivation of thrombin by heparin cofactor II. J Biol Chem, 279(11), 9785-95, 2004

Friedrich, R, Panizzi, P, Fuentes-Prior, P, Richter, K, Verhamme, I, Anderson, PJ, Kawabata, S, Huber, R, Bode, W, Bock, PE. Staphylocoagulase is a prototype for the mechanism of cofactor-induced zymogen activation. Nature, 425(6957), 535-9, 2003

Verhamme, IM, Olson, ST, Tollefsen, DM, Bock, PE. Binding of exosite ligands to human thrombin. Re-evaluation of allosteric linkage between thrombin exosites I and II. J Biol Chem, 277(9), 6788-98, 2002

Olson, ST, Swanson, R, Day, D, Verhamme, I, Kvassman, J, Shore, JD. Resolution of Michaelis complex, acylation, and conformational change steps in the reactions of the serpin, plasminogen activator inhibitor-1, with tissue plasminogen activator and trypsin. Biochemistry, 40(39), 11742-56, 2001

Boxrud, PD, Verhamme, IM, Fay, WP, Bock, PE. Streptokinase triggers conformational activation of plasminogen through specific interactions of the amino-terminal sequence and stabilizes the active zymogen conformation. J Biol Chem, 276(28), 26084-9, 2001

Verhamme, I, Kvassman, JO, Day, D, Debrock, S, Vleugels, N, Declerck, PJ, Shore, JD. Accelerated conversion of human plasminogen activator inhibitor-1 to its latent form by antibody binding. J Biol Chem, 274(25), 17511-7, 1999

Kvassman, JO, Verhamme, I, Shore, JD. Inhibitory mechanism of serpins: loop insertion forces acylation of plasminogen activator by plasminogen activator inhibitor-1. Biochemistry, 37(44), 15491-502, 1998

Gibson, A, Baburaj, K, Day, DE, Verhamme, I, Shore, JD, Peterson, CB. The use of fluorescent probes to characterize conformational changes in the interaction between vitronectin and plasminogen activator inhibitor-1. J Biol Chem, 272(8), 5112-21, 1997

Bock, PE, Day, DE, Verhamme, IM, Bernardo, MM, Olson, ST, Shore, JD. Analogs of human plasminogen that are labeled with fluorescence probes at the catalytic site of the zymogen. Preparation, characterization, and interaction with streptokinase. J Biol Chem, 271(2), 1072-80, 1996

Shore, JD, Day, DE, Francis-Chmura, AM, Verhamme, I, Kvassman, J, Lawrence, DA, Ginsburg, D. A fluorescent probe study of plasminogen activator inhibitor-1. Evidence for reactive center loop insertion and its role in the inhibitory mechanism. J Biol Chem, 270(10), 5395-8, 1995

Lawrence, DA, Ginsburg, D, Day, DE, Berkenpas, MB, Verhamme, IM, Kvassman, JO, Shore, JD. Serpin-protease complexes are trapped as stable acyl-enzyme intermediates. J Biol Chem, 270(43), 25309-12, 1995

Sherman, PM, Lawrence, DA, Verhamme, IM, Paielli, D, Shore, JD, Ginsburg, D. Identification of tissue-type plasminogen activator-specific plasminogen activator inhibitor-1 mutants. Evidence that second sites of interaction contribute to target specificity. J Biol Chem, 270(16), 9301-6, 1995

Den Tandt, WR, Inaba, T, Verhamme, I, Overdyk, B, Brouwer, J, Prieur, D. Non-identity of human plasma lysozyme and 4-methylumbelliferyl-tetra-N-acetyl-beta-D-chitotetraoside hydrolase. Int J Biochem, 20(7), 713-9, 1988

Verhamme, IM, Van Dedem, GW, Lauwers, AR. Ionic-strength-dependent substrate inhibition of the lysis of Micrococcus luteus by hen egg-white lysozyme. Eur J Biochem, 172(3), 615-20, 1988