Journal:Acta Cryst D:S2059798322007082

Native glycosylation and binding of the anti-depressant paroxetine in a low-resolution crystal structure of human myeloperoxidase

Lucas Krawczyk, Shubham Semwal, Jalal Soubhye, Salma Lemri Ouadriri, Martine Prévost, Pierre Van Antwerpen, Goedele Roos, & Julie Bouckaert ^[1]

Molecular Tour
The human enzyme myeloperoxidase (MPO in short) is a heme-dependent peroxidase. The peroxidases are a large group of enzymes playing roles in various biological processes. MPO is found in mammalian neutrophils, which is a type of with blood cells, and there it catalyzes the oxidation of halide ions and thiocyanate in the presence of hydrogen peroxide. This results in strong oxidizing agents and they are helping to eliminate the bacteria or viruses which were taken up by phagocytosis. So, MPO is a key protein in the human immune defense system.

In order for MPO to function correctly, sugar molecules need to be present on its protein surface. This is called the glycosylation pattern. Until now, only a part of the glycosylation of MPO could be seen from the crystal structures deposited in the Protein Data Bank, holding the collection of available structures of biomolecules. Human MPO has five glycosylation sites, identified at positions Asn323, Asn355, Asn391, Asn483 and Asn729. We present here a structure in which the glycan structures on these five glycosylation sites could be identified. and 30 glycan chains on the Asn-binding sites. Each chain has a different colour. The 8 monomers forming (each dimer has a different colour) in the crystal structure of human MPO, with their glycosylation structures as determined. (magenta), and each catalytic site carries an iron-containing (sea-green) and has an (CSO; in yellow) within 12 Å distance from the heme group. All starts with an N-acetylglucosamine (blue), modifying the labelled asparagine, and many are further substituted also with mannose (green) and fucose (red). Possible positions Asn323, Asn355, Asn391, Asn483 and Asn729 are labelled. Changes are possible due to N-glycan remodeling that occur post biogenesis and each glycosylation site may hold a truly individual protein function. For example, only the Asn323 site carries the peculiar phosphomannosylation, that it may be use to traffic into azurophilic granules. We compare these with the glycans identified in proteomics studies and from eighteen human MPO structures available in the PDB.

. Two N-glycans pack tightly with each other at the dimer interface. The heme groups are present in both monomers and are shown as a sea green spacefill model. Paroxetine (spacefill model) is bound in one monomer per dimer (chain H). Green spheres represent chloride ions, pink spheres are calcium ions and CPK compositions are phosphate ions. . Inter- and intermolecular hydrogen bonds, inclusive with water molecules (shown as red spheres), are shown as white dashed lines. , by symmetrically using N-acetylglucosamine (blue, middle) β1,2-linked to the α1,6 arm of the trimannose core (green) and the fucose (red) α1,6-linked to the core GlcNAc1.

carries an iron-containing heme group. The heme group of MPO and the near binding of the thiocyanate, a natural substrate, and paroxetine, an inhibitor, as found in the monomer with chain A are shown. Amino acid numbering is based on the pro-MPO crystal structure numbering, including signal and pro-peptide. Our structure also contains , a recently discovered inhibitor of MPO, previously used as anti-depressant. The bound paroxetine was always found in the presence of , a physiological substrate of MPO. A lot of effort has been undertaken into inhibitor design, as things can also go wrong with MPO. When things go wrong, MPO is released into the extracellular fluid. This circulating MPO damages host tissue as the reaction products of MPO can oxidize biomolecules (lipids, DNA and proteins). So, MPO is involved in a lot of pathologies, either as a source or to make the symptoms worse of existing pathologies, creating a large interest into the design of molecules in order to block this circulating MPO.

of PDB entry 1dnu (blue) containing thiocyanate (SCN-) with chain A of the MPO crystal structure (wheat), bound to SCN- and paroxetine.

References

1. ↑ Krawczyk L, Semwal S, Soubhye J, Lemri Ouadriri S, Prevost M, Van Antwerpen P, Roos G, Bouckaert J. Native glycosylation and binding of the antidepressant paroxetine in a low-resolution crystal structure of human myeloperoxidase. Acta Crystallogr D Struct Biol. 2022 Sep 1;78(Pt 9):1099-1109. doi:, 10.1107/S2059798322007082. Epub 2022 Aug 9. PMID:36048150 doi:http://dx.doi.org/10.1107/S2059798322007082