Human α-thrombin is a multifunctional serine protease that plays a pivotal role in hemostasis. Two binding sites on its surface, the fibrinogen recognition site (exosite I) and the heparin binding site (exosite II), control its catalytic function. Promising thrombin inhibitors are aptamers, short single-stranded oligonucleotides able to bind target proteins with high selectivity and affinity (Bock et al, Nature, 1992, 355, 564-6). Among them a 15-mer aptamer recognizing exosite I (TBA) (Macaya et al, PNAS, 1993, 90, 3745-9) and a 29-mer binding exosite II (HD22) aptamer (Tasset et al, JMB, 1997, 272, 688-98) were selected for their high affinity towards thrombin. Structural data on thrombin-aptamer complexes are essential to understand recognition mechanism(s) of TBA and HD22 and to design novel aptamers with improved pharmaceutical properties. We have performed an X-ray structural characterization of the complexes of thrombin with TBA (Russo Krauss et al, NAR, 2012, 40, 8119-28), TBA variants (Russo Krauss et al, NAR, 2011, 39, 7858-67) and mutants or a shorter variant of HD22 (HD22-27mer). This study has been complemented with a CD analysis of aptamers both free and in complex with thrombin, in the presence of Na+ or K+. All together our results have provided a clear picture of the recognition process between thrombin and its aptamers. Moreover, they allow to define the effect that ions have on structure and flexibility, and therefore on thrombin affinity and inhibition, of the various aptamers. Notably, the thrombin-HD22-27mer complex has provided the first crystallographic model of an aptamer with a mixed duplex-quadruplex fold and reveals unexpected novel structural features.

Structural motifs that govern the recognition process between thrombin and aptamers inhibiting exosite I or II / RUSSO KRAUSS, Irene; Pica, Andrea; Merlino, Antonello; L., Mazzarella; Sica, Filomena. - STAMPA. - (2013), pp. P36-P36. (Intervento presentato al convegno IV International Meeting on G-quadruplex Nucleic Acids from Structure to Chemistry and Biology tenutosi a Nanyang Executive Center, Nanyang Technological University, Singapore nel 1-4 Luglio 2013).

Structural motifs that govern the recognition process between thrombin and aptamers inhibiting exosite I or II

RUSSO KRAUSS, IRENE;PICA, ANDREA;MERLINO, ANTONELLO;SICA, FILOMENA
2013

Abstract

Human α-thrombin is a multifunctional serine protease that plays a pivotal role in hemostasis. Two binding sites on its surface, the fibrinogen recognition site (exosite I) and the heparin binding site (exosite II), control its catalytic function. Promising thrombin inhibitors are aptamers, short single-stranded oligonucleotides able to bind target proteins with high selectivity and affinity (Bock et al, Nature, 1992, 355, 564-6). Among them a 15-mer aptamer recognizing exosite I (TBA) (Macaya et al, PNAS, 1993, 90, 3745-9) and a 29-mer binding exosite II (HD22) aptamer (Tasset et al, JMB, 1997, 272, 688-98) were selected for their high affinity towards thrombin. Structural data on thrombin-aptamer complexes are essential to understand recognition mechanism(s) of TBA and HD22 and to design novel aptamers with improved pharmaceutical properties. We have performed an X-ray structural characterization of the complexes of thrombin with TBA (Russo Krauss et al, NAR, 2012, 40, 8119-28), TBA variants (Russo Krauss et al, NAR, 2011, 39, 7858-67) and mutants or a shorter variant of HD22 (HD22-27mer). This study has been complemented with a CD analysis of aptamers both free and in complex with thrombin, in the presence of Na+ or K+. All together our results have provided a clear picture of the recognition process between thrombin and its aptamers. Moreover, they allow to define the effect that ions have on structure and flexibility, and therefore on thrombin affinity and inhibition, of the various aptamers. Notably, the thrombin-HD22-27mer complex has provided the first crystallographic model of an aptamer with a mixed duplex-quadruplex fold and reveals unexpected novel structural features.
2013
Structural motifs that govern the recognition process between thrombin and aptamers inhibiting exosite I or II / RUSSO KRAUSS, Irene; Pica, Andrea; Merlino, Antonello; L., Mazzarella; Sica, Filomena. - STAMPA. - (2013), pp. P36-P36. (Intervento presentato al convegno IV International Meeting on G-quadruplex Nucleic Acids from Structure to Chemistry and Biology tenutosi a Nanyang Executive Center, Nanyang Technological University, Singapore nel 1-4 Luglio 2013).
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11588/563678
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