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Recognition and Conformational Properties of an Alternative Antithrombin Binding Sequence Obtained by Chemoenzymatic Synthesis.

ABSTRACT: Heparin is a highly sulfated glycosaminoglycan (GAG) of natural origin used as an anticoagulant and antithrombotic drug. These properties are principally based on the binding and activation of antithrombin (AT) through the pentasaccharide sequence GlcNAc/NS,6S-GlcA-GlcNS,3,6S-IdoA2S-GlcNS,6S (AGA*IA). Literature data show that the population of the 2 S0 ring conformation of the 2-O-sulfo-?-l-iduronic acid (IdoA2S) motif correlates with the affinity and activation of AT. It was recently demonstrated that two synthetic AGA*IA-containing hexasaccharides (one G unit added at the reducing end), differing in the degree of sulfation of the IdoA unit, show comparable affinity and ability to activate AT, despite a different conformation of the IdoA residue. In this paper, the binding of these two glycans to AT was studied by isothermal titration microcalorimetry (ITC), transferred (tr-) NOESY, saturation transfer difference (STD) NMR spectroscopy and molecular dynamics (MD) simulations. Results indicated that both the IdoA2S and the IdoA units assume a 2 S0 conformation when bound with AT, and so present a common binding epitope for the two glycans, centred on the AGA*IA sequence.

SUBMITTER: Stancanelli E

PROVIDER: S-EPMC6517080 | biostudies-literature | 2018 Mar

REPOSITORIES: biostudies-literature

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Recognition and Conformational Properties of an Alternative Antithrombin Binding Sequence Obtained by Chemoenzymatic Synthesis.

Stancanelli Eduardo E Elli Stefano S Hsieh Po-Hung PH Liu Jian J Guerrini Marco M

Chembiochem : a European journal of chemical biology 20180324

Heparin is a highly sulfated glycosaminoglycan (GAG) of natural origin used as an anticoagulant and antithrombotic drug. These properties are principally based on the binding and activation of antithrombin (AT) through the pentasaccharide sequence GlcNAc/NS,6S-GlcA-GlcNS,3,6S-IdoA2S-GlcNS,6S (AGA*IA). Literature data show that the population of the <sup>2</sup> S<sub>0</sub> ring conformation of the 2-O-sulfo-α-l-iduronic acid (IdoA2S) motif correlates with the affinity and activation of AT. It ...[more]

PMID: 29573524

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Project description:The present study deals with the conformation in solution of two heparin octasaccharides containing the pentasaccharide sequence GlcN(NAc,6S)-GlcA-GlcN(NS,3,6S)-IdoA(2S)-GlcN(NS,6S) [AGA*IA; where GlcN(NAc,6S) is N-acetylated, 6-O-sulfated alpha-D-glucosamine, GlcN(NS,3,6S) is N,3,6-O-trisulfated alpha-D-glucosamine and IdoA(2S) is 2-O-sulfated IdoA (alpha-L-iduronic acid)] located at different positions in the heparin chain and focuses on establishing geometries of IdoA residues (IdoA(2S) and IdoA) both inside and outside the AGA*IA sequence. AGA*IA constitutes the active site for AT (antithrombin) and is essential for the expression of high anticoagulant and antithrombotic activities. Analysis of NMR parameters [NOEs (nuclear Overhauser effects), transferred NOEs and coupling constants] for the two octasaccharides indicated that between the 1C4 and 2S0 conformations present in dynamic equilibrium in the free state for the IdoA(2S) residue within AGA*IA, AT selects the 2S0 form, as previously shown [Hricovini, Guerrini, Bisio, Torri, Petitou and Casu (2001) Biochem. J. 359, 265-272]. Notably, the 2S0 conformation is also adopted by the non-sulfated IdoA residue preceding AGA*IA that, in the absence of AT, adopts predominantly the 1C4 form. These results further support the concept that heparin-binding proteins influence the conformational equilibrium of iduronic acid residues that are directly or indirectly involved in binding and select one of their equi-energetic conformations for best fitting in the complex. The complete reversal of an iduronic acid conformation preferred in the free state is also demonstrated for the first time. Preliminary docking studies provided information on the octasaccharide binding location agreeing most closely with the experimental data. These results suggest a possible biological role for the non-sulfated IdoA residue preceding AGA*IA, previously thought not to influence the AT-binding properties of the pentasaccharide. Thus, for each AT binding sequence longer than AGA*IA, the interactions with the protein could differ and give to each heparin fragment a specific biological response.

Dataset Information

Recognition and Conformational Properties of an Alternative Antithrombin Binding Sequence Obtained by Chemoenzymatic Synthesis.

Publications

Recognition and Conformational Properties of an Alternative Antithrombin Binding Sequence Obtained by Chemoenzymatic Synthesis.

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