Structure of Human Chitotriosidase

Abstract

Chitin hydrolases have been identified in a variety of organisms ranging from bacteria to eukaryotes. They have been proposed to be possible targets for the design of novel chemotherapeutics against human pathogens such as fungi and protozoan parasites as mammals were not thought to possess chitin-processing enzymes. Recently, a human chitotriosidase was described as a marker for Gaucher disease with plasma levels of the enzyme elevated up to 2 orders of magnitude. The chitotriosidase was shown to be active against colloidal chitin and is inhibited by the family 18 chitinase inhibitor allosamidin. Here, the crystal structure of the human chitotriosidase and complexes with a chitooligosaccharide and allosamidin are described. The structures reveal an elongated active site cleft, compatible with the binding of long chitin polymers, and explain the inactivation of the enzyme through an inherited genetic deficiency. Comparison with YM1 and HCgp-39 shows how the chitinase has evolved into these mammalian lectins by the mutation of key residues in the active site, tuning the substrate binding specificity. The soaking experiments with allosamidin and chitooligosaccharides give insight into ligand binding properties and allow the evaluation of differential binding and design of speciesselective chitinase inhibitors.

Crystal Structures of Allosamidin Derivatives in Complex with Human Macrophage Chitinase

“…As reported earlier, soaking of HCHT crystals with ALLO and its derivatives resulted in severe cracking (21). To overcome these problems, HCHT was co-crystallized with ALLO and its derivatives DEME, METH, and GLCB (Fig.…”

“…Structure Determination-Human chitinase (HCHT) was isolated as described previously (21). As reported earlier, soaking of HCHT crystals with ALLO and its derivatives resulted in severe cracking (21).…”

“…Data were collected on beamline ID14-EH2 at the European Synchrotron Radiation Facility (Grenoble, France) and beamline X11 at the Deutsches Elektronen Synchrotron (the Deutsches Elektronen Synchrotron, Hamburg, Germany), and processed with the HKL suite of programs (32) ( Table II). The HCHT⅐ALLO structure was solved by molecular replacement with AMoRe (33) (search model, the native HCHT structure (21); top solution, r ϭ 0.344; correlation coefficient, 0.694) and was used as a starting structure for the refinement of the other complexes. Refinement was performed with CNS (34) interspersed with model building in O (35).…”

“…A preliminary soaking study has also been reported for the human chitinase (21). The inhibitor appears to bind from the Ϫ3 to Ϫ1 subsites, with the allosamizoline occupying the Ϫ1 subsite.…”

Screening-based Discovery and Structural Dissection of a Novel Family 18 Chitinase Inhibitor

“…Thr 138 and Tyr 139 form one wall of the subsite binding the secondary caffeine, and whereas their side chains do not form hydrogen bonds with the ligand, they make extensive van der Waals interactions that are likely to contribute significantly to ligand binding and stabilization. Crystal structures of HCHT (17,34) show that the asparagine replacing Thr 138 adopts a conformation similar to that of the threonine and thus is unlikely to significantly alter binding. On the other hand, the side chain of the phenylalanine replacing Tyr 139 points away from the active site, into a pocket created by a glycine in place of AfChiB1 Asn 78 .…”