Unexpected Active-Site Flexibility in the Structure of Human Neutrophil Elastase in Complex with a New Dihydropyrimidone Inhibitor

Abstract

Human neutrophil elastase (HNE), a trypsin-type serine protease, is of pivotal importance in the onset and progression of chronic obstructive pulmonary disease (COPD). COPD encompasses a group of slowly progressive respiratory disorders and is a major medical problem and the fifth leading cause of death worldwide. HNE is a major target for the development of compounds that inhibit the progression of long-term lung function decline in COPD patients. Here, we present the three-dimensional structure of a potent dihydropyrimidone inhibitor (DHPI) non-covalently bound to HNE at a resolution of 2.0 Å. The inhibitor binds to the active site in a unique orientation addressing S1 and S2 subsites of the protease. To facilitate further analysis of this binding mode, we determined the structure of the uncomplexed enzyme at a resolution of 1.86 Å. Detailed comparisons of the HNE:DHPI complex with the uncomplexed HNE structure and published structures of other elastase:inhibitor complexes revealed that binding of DHPI leads to large conformational changes in residues located in the S2 subsite. The rearrangement of residues Asp95-Leu99B creates a deep, well-defined cavity, which is filled by the P2 moiety of the inhibitor molecule to almost perfect shape complementarity. The shape of the S2 subsite in complex with DHPI clearly differs from all other observed HNE structures. The observed structural flexibility of the S2 subsite is a key feature for the understanding of the binding mode of DHPIs in general and the development of new HNE selective inhibitors.

Neutrophil elastase inhibitors: a patent review and potential applications for inflammatory lung diseases (2010 – 2014)

“…The dihydropyrimidinones function as calcium channel modulators and selective a1a-adrenoceptor antagonists. Recently, derivatives of these compounds have been found to be potent human NE inhibitors and have thus been proposed as potential COPD therapeutics [66]. Twenty-nine 4-(4-cyano-2-thioaryl)-dihydropyrimidinone (Table 3) derivatives (19) and 273 other substituted bicyclic dihydropyrimidinone derivatives (20) claimed in reviewed patents potently inhibit NE, and the IC 50 values for most of these compounds are in the picomolar to low nanomolar range [67].…”

Inhibitors and Antibody Fragments as Potential Anti-Inflammatory Therapeutics Targeting Neutrophil Proteinase 3 in Human Disease

“…Several threedimensional structures of HNE complexed with the third domain of turkey ovomucoid (Bode et al, 1986), domain 2 of SLPI (Koizumi et al, 2008), and synthetic inhibitors have all been determined (Wei et al, 1988;Navia et al, 1989;Huang et al, 2008;Hansen et al, 2011;Lechtenberg et al, 2015). PR3 and HNE share a high sequence identity (56%) and display a common fold.…”

Discovery of C-shaped aurone human neutrophil elastase inhibitors

“…One of the main pitfalls of the drug discovery process toward HNErelated diseases is the target flexibility due to induced-fit events upon the binding of different ligands. 3,4 For example a potent noncovalent dihydropyrimidone inhibitor was co-crystallised with HNE revealing a unique orientation addressing HNE S1 and S2 subsites, in which the formation of a deep and well-defined cavity at S2 was first described (Fig. 1).…”