Bivalent inhibition of human beta-tryptase

Background: Human beta -tryptase is a mast cell specific trypsinlike serine protease that is thought to play a key role in the pathogenesis of diverse allergic and inflammatory disorders like asthma and psoriasis. The recentl y resolved crystal structure revealed that the enzymatically active tetrame r consists of four quasi-identical monomers. The spatial display of the fou r identical active sites represents an ideal basis for the rational design of bivalent inhibitors. Results: Based on modeling experiments homobivalent inhibitors were constru cted using (i) 6A,6D-dideoxy-6A,6D-diamino-beta -cyclodextrin as a rigid te mplate to bridge the space between the two pairs of identical active sites and (ii) 3-(aminomethyl)benzene as a headgroup to occupy the arginine/lysin e specific S1 subsites. A comparative analysis of the inhibitory potencies of synthetic constructs that differ in size and type of the spacer between headgroup and template revealed that the construct contained two 3-(aminome thyl)benzenesulfonyl-glycine groups linked to the 6A,6D-diamino groups of b eta -cyclodextrin as an almost ideal bivalent inhibitor with a cooperativit y factor of 1.9 vs. the ideal value of 2. The bivalent binding mode is supp orted by the inhibitor/tetramer ratio of 2:1 required for inactivation of t ryptase and by X-ray analysis of the inhibitor/tryptase complex. Conclusion: The results obtained with the rigid cyclodextrin template under lined the importance of a minimal loss of conformational entropy in bivalen t binding, but also showed the limitations imposed by such rigid core molec ules in terms of optimal occupancy of binding sites and thus of enthalpic s trains in bidentate binding modes. The main advantage of bivalent inhibitor s is their high selectivity for the target enzyme that can be achieved util izing the principle of multivalency. (C) 2001 Elsevier Science Ltd. All rig hts reserved.