title: Human Chitotriosidase Is an Endo-Processive Enzyme.
abstract: Human chitotriosidase (HCHT) is involved in immune response to chitin-containing pathogens in humans. The enzyme is able to degrade chitooligosaccharides as well as crystalline chitin. The catalytic domain of HCHT is connected to the carbohydrate binding module (CBM) through a flexible hinge region. In humans, two active isoforms of HCHT are found-the full length enzyme and its truncated version lacking CBM and the hinge region. The active site architecture of HCHT is reminiscent to that of the reducing-end exo-acting processive chitinase ChiA from bacterium Serratia marcescens (SmChiA). However, the presence of flexible hinge region and occurrence of two active isoforms are reminiscent to that of non-processive endo-chitinase from S. marcescens, SmChiC. Although the studies on soluble chitin derivatives suggest the endo-character of HCHT, the mode of action of the enzyme on crystalline chitin is not known. Here, we made a thorough characterization of HCHT in terms of the mode of action, processivity, binding, and rate constants for the catalysis and dissociation using α-chitin as substrate. HCHT efficiently released the end-label from reducing-end labelled chitin and had also high probability (95%) of endo-mode initiation of processive run. These results qualify HCHT as an endo-processive enzyme. Processivity and the rate constant of dissociation of HCHT were found to be in-between those, characteristic to processive exo-enzymes, like SmChiA and randomly acting non-processive endo-enzymes, like SmChiC. Apart from increasing the affinity for chitin, CBM had no major effect on kinetic properties of HCHT.
authors.affiliation: Institute of Molecular and Cell Biology, University of Tartu, Tartu, Estonia.
authors.forename: Silja
authors.identifier: http://orcid.org/0000-0001-7018-4122
authors.initials: S
authors.lastname: Kuusk
authors.affiliation: Department of Chemistry, Biotechnology and Food Science, Norwegian University of Life Sciences, Ås, Norway.
authors.forename: Morten
authors.initials: M
authors.lastname: Sørlie
authors.affiliation: Institute of Molecular and Cell Biology, University of Tartu, Tartu, Estonia.
authors.forename: Priit
authors.initials: P
authors.lastname: Väljamäe
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics; D002384:Catalysis; D020134:Catalytic Domain; D002686:Chitin / Q000031:analogs & derivatives / Q000737:chemistry*; D002688:Chitinases / Q000737:chemistry* / Q000235:genetics; D048271:Chitosan / Q000737:chemistry*; D006596:Hexosaminidases / Q000737:chemistry* / Q000235:genetics; D006801:Humans; D007113:Immunity, Innate / Q000235:genetics; D007700:Kinetics; D009844:Oligosaccharides; D012706:Serratia marcescens / Q000201:enzymology; D013379:Substrate Specificity
chemical_list: D001426:Bacterial Proteins; D009844:Oligosaccharides; C493484:oligochitosan; D002686:Chitin; D048271:Chitosan; D006596:Hexosaminidases; C086400:chitotriosidase; D002688:Chitinases; C000601656:chitinase A, Serratia marcescens