Abstract
Family 18 chitinases play key roles in the life cycles of a variety of organisms ranging from bacteria to man. Very recently it has been shown that one of the mammalian chitinases is highly overexpressed in the asthmatic lung and contributes to the pathogenic process through recruitment of inflammatory cells. Although several potent natural product chitinase inhibitors have been identified, their chemotherapeutic potential or their use as cell biological tools is limited due to their size, complex chemistry, and limited availability. We describe a virtual screening-based approach to identification of a novel, purine-based, chitinase inhibitor. This inhibitor acts in the low micromolar (Ki=2.8+/-0.2 microM) range in a competitive mode. Dissection of the binding mode by x-ray crystallography reveals that the compound, which consists of two linked caffeine moieties, binds in the active site through extensive and not previously observed stacking interactions with conserved, solvent exposed tryptophans. Such exposed aromatics are also present in the structures of many other carbohydrate processing enzymes. The compound exhibits favorable chemical properties and is likely to be useful as a general scaffold for development of pan-family 18 chitinase inhibitors.
Highlights
18 chitinases (CAZy GH 18) hydrolyze chitin, a homopolymer of -(1,4)-linked N-acetylglucosamine
Chitin is a key component of the fungal cell wall, and chitinases are thought to be required for fungal cell separation and morphogenesis [2,3,4]
Whereas the method identifies both known and novel A. fumigatus chitinase B1 (AfChiB1) inhibitors, a large number of predicted hits turn out to be false positives, which is unexpected, in particular as the assumptions made to speed up the virtual screening should, if anything, give rise to false negative results
Summary
18 chitinases (CAZy GH 18) hydrolyze chitin, a homopolymer of -(1,4)-linked N-acetylglucosamine. Whereas chitin does not occur in humans, it is known that we possess two chitinases [5, 6]. A high throughput screen identified three xanthine derivatives, theophylline, caffeine, and pentoxifylline, as competitive inhibitors of bacterial, fungal, and human family 18 chitinases [19]. Their inhibition was not strong (high micromolar), analysis of their mode of inhibition revealed that these molecules mimic many of the chitinase-substrate hydrogen bonds and in addition display extensive - stacking interactions with a conserved tryptophan. The virtual screen resulted in the identification of a new xanthine-based chitinase ligand that competitively inhibits several family 18 chitinases in the low micromolar range
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