Dependence of salt concentration on glycosaminoglycan-lysozyme interactions in cartilage.
The cationic protein, lysozyme, has an extracellular distribution in cartilage but its precise role in this tissue has not yet been established. This study describes the dependence of salt concentration on the binding properties of lysozyme isoforms of different cationic charges, isolated from bovine cartilage, to the two major and structurally similar glycosaminoglycans of cartilage, i.e., chondroitin sulfate and hyaluronan. The binding of most cartilage lysozyme isoforms and hen egg-white lysozyme (control) to chondroitin sulfate and hyaluronan linked to agarose supports displayed optimal levels at approximately 20 and 5-10 mM salt, respectively, but decreased at both lower and higher salt concentrations indicating the electrostatic nature of the interactions. However, optimal binding of the most cationic lysozyme isoform to chondroitin sulfate occurred at 60 mM salt, with significant binding remaining at 150 mM. This isoform also showed binding to hyaluronan up to 60 mM salt, while for the other isoforms binding was observed only up to 150 and 40 mM salt for chondroitin sulfate and hyaluronan, respectively. The low salt concentrations at which these interactions occur are likely to exist in cartilage as shown from equilibrium dialysis studies performed using solutions of chondroitin sulfate (up to 10%, a concentration likely to occur in cartilage). From Scatchard analysis, the affinity of binding of all lysozymes to chondroitin sulfate was similar (Kd = 10(-6) M) and slightly lower than their binding to hyaluronan (Kd = 10(-7) M) of similar molecular mass.
Moss
J M
JM
Department of Biochemistry and Molecular Biology, Monash University, Clayton, Victoria, Australia.
Van Damme
M P
MP
Murphy
W H
WH
Preston
B N
BN
D000818:Animals; D002358:Cartilage, Articular / Q000201:enzymology*; D002417:Cattle; D002809:Chondroitin Sulfates / Q000378:metabolism*; D006025:Glycosaminoglycans / Q000737:chemistry / Q000378:metabolism*; D006820:Hyaluronic Acid / Q000378:metabolism*; D007700:Kinetics; D009113:Muramidase / Q000737:chemistry / Q000378:metabolism*; D009300:Nasal Septum / Q000201:enzymology*; D009994:Osmolar Concentration; D011485:Protein Binding; D012685:Sepharose
D006025:Glycosaminoglycans; D006820:Hyaluronic Acid; D002809:Chondroitin Sulfates; D012685:Sepharose; D009113:Muramidase