Effect of body temperature on chondroitinase ABC's ability to cleave chondroitin sulfate glycosaminoglycans.
Chondroitinase ABC (Ch'ase ABC) is a bacterial lyase that degrades chondroitin sulfate (CS), dermatan sulfate, and hyaluronan glycosaminoglycans (GAGs). This enzyme has received significant attention as a potential therapy for promoting central nervous system and peripheral nervous system repair based on its degradation of CS GAGs. Determination of the stability of Ch'ase ABC activity at temperatures equivalent to normal (37 degrees C) and elevated (39 degrees C) body temperatures is important for optimizing its clinical usage. We report here data obtained from examining enzymatic activity at these temperatures across nine lots of commercially available protease-free Ch'ase ABC. CS GAG degrading activity was assayed by using 1) immunohistochemical detection of unsaturated disaccharide stubs generated by digestion of proteoglycans in tissue sections and 2) fluorophore-assisted carbohydrate electrophoresis (FACE) and/or high-performance liquid chromatography (HPLC) to separate and quantify unsaturated disaccharide digestion products. Our results indicate that there is a significant effect of lot and time on enzymatic thermostability. Average enzymatic activity is significantly decreased at 1 and 3 days at 39 degrees C and 37 degrees C, respectively. Furthermore, the average activity seen after 1 day was significantly different between the two temperatures. Addition of bovine serum albumin as a stabilizer significantly preserved enzymatic activity at 1 day, but not 3 days, at 39 degrees C. These results show that the CS GAG degrading activity of Ch'ase ABC is significantly decreased with incubation at body temperature over time and that all lots do not show equal thermostability. These findings are important for the design and interpretation of experimental and potential clinical studies involving Ch'ase ABC.
Tester
Nicole J
NJ
Malcom Randall Veterans Affairs Medical Center, Gainesville, Florida, USA.
Plaas
Anna H
AH
Howland
Dena R
DR
D000818:Animals; D001831:Body Temperature / Q000502:physiology*; D002415:Cats; D002417:Cattle; D019765:Chondroitin ABC Lyase / Q000737:chemistry* / Q000378:metabolism*; D002809:Chondroitin Sulfates / Q000378:metabolism*; D004789:Enzyme Activation / Q000502:physiology; D004795:Enzyme Stability / Q000502:physiology; D006801:Humans; D051381:Rats; D020318:Rats, Long-Evans; D012710:Serum Albumin, Bovine / Q000737:chemistry / Q000494:pharmacology; D013997:Time Factors
C038142:chondroitin sulfate glycosaminoglycan; D012710:Serum Albumin, Bovine; D002809:Chondroitin Sulfates; D019765:Chondroitin ABC Lyase