A dynamic Asp–Arg interaction is essential for catalysis in microsomal prostaglandin E₂ synthase

Microsomal prostaglandin E₂ synthase type 1 (mPGES-1) is responsible for the formation of the potent lipid mediator prostaglandin E₂ under proinflammatory conditions, and this enzyme has received considerable attention as a drug target. Recently, a high-resolution crystal structure of human mPGES-1...

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Veröffentlicht in:Proceedings of the National Academy of Sciences - PNAS 2016-01, Vol.113 (4), p.972-977
Hauptverfasser: Brock, Joseph S., Hamberg, Mats, Balagunaseelan, Navisraj, Goodman, Michael, Morgenstern, Ralf, Strandback, Emilia, Samuelsson, Bengt, Rinaldo-Matthis, Agnes, Haeggström, Jesper Z.
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Sprache:eng
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Zusammenfassung:Microsomal prostaglandin E₂ synthase type 1 (mPGES-1) is responsible for the formation of the potent lipid mediator prostaglandin E₂ under proinflammatory conditions, and this enzyme has received considerable attention as a drug target. Recently, a high-resolution crystal structure of human mPGES-1 was presented, with Ser-127 being proposed as the hydrogen-bond donor stabilizing thiolate anion formation within the cofactor, glutathione (GSH). We have combined site-directed mutagenesis and activity assays with a structural dynamics analysis to probe the functional roles of such putative catalytic residues. We found that Ser-127 is not required for activity, whereas an interaction between Arg-126 and Asp-49 is essential for catalysis. We postulate that both residues, in addition to a crystallographic water, serve critical roles within the enzymatic mechanism. After characterizing the size or charge conservative mutations Arg-126–Gln, Asp-49–Asn, and Arg-126–Lys, we inferred that a crystallographic water acts as a general base during GSH thiolate formation, stabilized by interaction with Arg-126, which is itself modulated by its respective interaction with Asp-49. We subsequently found hidden conformational ensembles within the crystal structure that correlate well with our biochemical data. The resulting contact signaling network connects Asp-49 to distal residues involved in GSH binding and is ligand dependent. Our work has broad implications for development of efficient mPGES-1 inhibitors, potential anti-inflammatory and anticancer agents.
ISSN:0027-8424
1091-6490
1091-6490
DOI:10.1073/pnas.1522891113