Development of transition state analogues targeting chitinases and oligosaccharyl transferase

Abstract

Oligosaccharyl transferase (OT) plays a central role in the biosynthesis of asparagine-linked glycoproteins in eukaryotic systems. The glycosylation step catalyzed by OT involves the co-translational transfer of a tetradecasaccharide from a dolichyl-pyrophosphate carrier to an asparagine side-chain within the Asn-Xaa-Ser/Thr sequence of a nascent polypeptide. Chitinases, which was emerged as a therapeutic target in combating asthma, are [beta]-1,4-N-acetylglucosaminidases that hydrolyze chitin to generate the disaccharide chitobiose Although the reactions catalyzed by the two enzymes follow different pathways, they are believed to share similar transition states involving an oxocarbenium ion. To understand the mechanism of OT and discover potent and selective inhibitors against different chitinases, our intent was to utilize the common transition state analogue for both enzymes and systematically introduce additional binding determinants. The pseudo-disaccharides containing an imino sugar were designed to target the oxocarbenium ion like transition state. The pseudo-disaccharides containing imino sugar were synthesized and evaluated at inhibitors for OT and chitinases.

(cont.) Highlights and supporting studies from this work include: (1) the use of the Amadori rearrangement to generate the acyclic substrate; (2) the glycosylation of [beta]-hydroxy ketone; (3) the intramolecular reductive amination between the in-situ generated amine from azido and ketone moieties; (4) the determination of the stereo-chemical outcome by NOE difference experiments. The pseudo-disaccharides containing imino sugars exhibited IC50s in the low micromolar range versus chitinase, yet significant inhibitory activity against OT was not observed.