Human YKL-39 is a pseudo-chitinase with retained chitooligosaccharide-binding properties

Abstract

The chitinase-like proteins YKL-39 (chitinase 3-like-2) and YKL-40 (chitinase 3-like-1) are highly expressed in a number of human cells independent of their origin (mesenchymal, epithelial or haemapoietic). Elevated serum levels of YKL-40 have been associated with a negative outcome in a number of diseases ranging from cancer to inflammation and asthma. YKL-39 expression has been associated with osteoarthritis. However, despite the reported association with disease, the physiological or pathological role of these proteins is still very poorly understood. Although YKL-39 is homologous to the two family 18 chitinases in the human genome, it has been reported to lack any chitinase activity. In the present study, we show that human YKL-39 possesses a chitinase-like fold, but lacks key active-site residues required for catalysis. A glycan screen identified oligomers of N-acetylglucosamine as preferred binding partners. YKL-39 binds chitooligosaccharides and a newly synthesized derivative of the bisdionin chitinase-inhibitor class with micromolar affinity, through a number of conserved tryptophan residues. Strikingly, the chitinase activity of YKL-39 was recovered by reverting two non-conservative substitutions in the active site to those found in the active enzymes, suggesting that YKL-39 is a pseudo-chitinase with retention of chitinase-like ligand-binding properties.

Structural and Thermodynamic Insights into Chitooligosaccharide Binding to Human Cartilage Chitinase 3-like Protein 2 (CHI3L2 or YKL-39)

“…MembersSuperimposition of the structure of the wild-type YKL39⅐ GlcNAc) 6 complex obtained in this study on that of YKL39⅐GlcNAc 6 complex (4AY1) reported by Schimpl et al (27) results in a root mean square deviation of 0.22 for C␣ positions over 334 atoms (Fig. 5).…”

“…All diffraction data were indexed, integrated, and scaled using the program HKL2000 (29), and molecular replacement was employed to obtain phase information using the program MOLREP from the CCP4 suite (30). The structure of YKL-39 bound to GlcNAc 2 was solved using the previously published structure of YKL-39 in complex with GlcNAc 6 (PDB code 4AY1) as the search model (27). Other data sets, including ligand-free YKL-39 and YKL-39 in complex with GlcNAc 4 and GlcNAc 6 , were solved using the final structure of the YKL-39⅐GlcNAc 2 complex as the model for rigid body refinement.…”

“…In the structure by Schimpl et al (27), Asn-35 was mutated to Gln to prevent glycosylation, generating mutant N35Q. In this study, YKL-39 was functionally expressed in E. coli as a native, nonglycosylated protein.…”

“…Previously, the crystal structure of an N35Q mutant of YKL-39, bound to GlcNAc 6 , was reported (27). In the work reported here, we employ both crystallographic and thermodynamic studies to evaluate the binding of chitooligosaccharides of different lengths to YKL-39.…”

Periplasmic de-acylase helps bacteria don their biofilm coat

“…Such seemingly inactive glycoside hydrolase mutants in which catalytic residues have been replaced are an increasingly uncovered phenomenon, particularly in the case of chitinases, and generally thought to represent simple binding domains (10,11). Indeed, through fluorescence perturbation studies, the authors (5) measure weak but progressively tighter binding of β-1,6-GlcNAc oligomers of increasing size, whereas chito-oligosaccharides are shown not to bind.…”