Loop Electrostatics Asymmetry Modulates the Preexisting Conformational Equilibrium in Thrombin

Pozzi, Nicola; Zerbetto, Mirco; Acquasaliente, Laura; Tescari, Simone; Frezzato, Diego; Polimeno, Antonino; Gohara, David W.; Di Cera, Enrico; De Filippis, Vincenzo

doi:10.1021/acs.biochem.6b00385

Title: Loop Electrostatics Asymmetry Modulates the Preexisting Conformational Equilibrium in Thrombin

Journal Article · Mon Jun 27 00:00:00 EDT 2016 · Biochemistry

DOI:https://doi.org/10.1021/acs.biochem.6b00385· OSTI ID:1274760

Pozzi, Nicola ^[1]; Zerbetto, Mirco ^[2]; Acquasaliente, Laura ^[2]; Tescari, Simone ^[2]; Frezzato, Diego ^[2]; Polimeno, Antonino ^[2]; Gohara, David W. ^[1]; Di Cera, Enrico ^[1]; De Filippis, Vincenzo ^[2]

Saint Louis Univ. School of Medicine, St. Louis, MO (United States)
Univ. of Padua (Italy)

Thrombin exists as an ensemble of active (E) and inactive (E*) conformations that differ in their accessibility to the active site. Here we show that redistribution of the E*–E equilibrium can be achieved by perturbing the electrostatic properties of the enzyme. Removal of the negative charge of the catalytic Asp102 or Asp189 in the primary specificity site destabilizes the E form and causes a shift in the 215–217 segment that compromises substrate entrance. Solution studies and existing structures of D102N document stabilization of the E* form. A new high-resolution structure of D189A also reveals the mutant in the collapsed E* form. Furthermore, these findings establish a new paradigm for the control of the E*–E equilibrium in the trypsin fold.

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Research Organization:: Argonne National Laboratory (ANL), Argonne, IL (United States)

Sponsoring Organization:: National Inst. of Health

Grant/Contract Number:: HL049413; HL073813; HL112303

OSTI ID:: 1274760

Journal Information:: Biochemistry, Vol. 55, Issue 28; ISSN 0006-2960

Publisher:: American Chemical Society (ACS)Copyright Statement

Country of Publication:: United States

Language:: ENGLISH

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Cited by: 15 works

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Title: Loop Electrostatics Asymmetry Modulates the Preexisting Conformational Equilibrium in Thrombin

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