Two distinct chondroitin sulfate ABC lyases. An endoeliminase yielding tetrasaccharides and an exoeliminase preferentially acting on oligosaccharides.
Crude enzyme obtained from chondroitin sulfate-induced Proteus vulgaris NCTC 4636 has been fractionated into 1) an endoeliminase capable of depolymerizing chondroitin sulfate and related polysaccharides to produce, as end products, a mixture of Delta4-unsaturated tetra- and disaccharides and 2) an exoeliminase preferentially acting on chondroitin sulfate tetra- and hexasaccharides to yield the respective disaccharides. Isolation of the two enzymes was achieved by a simple two-step procedure: extracting the enzymes from intact P. vulgaris cells with a buffer solution of nonionic surfactant and then treating the extract by cation-exchange chromatography. Each of the enzymes thus prepared was apparently homogeneous as assessed by SDS-polyacrylamide gel electrophoresis and readily crystallized from polyethylene glycol solutions. Both enzymes acted on various substrates such as chondroitin sulfate, chondroitin sulfate proteoglycan, and dermatan sulfate at high, but significantly different, initial rates. They also attacked hyaluronan but at far lower rates and were inactive to keratan sulfate, heparan sulfate, and heparin. Our results show that the known ability of the conventional enzyme called "chondroitinase ABC" to catalyze the complete depolymerization of chondroitin sulfates to unsaturated disaccharides may actually result from the combination reactions by endoeliminase (chondroitin sulfate ABC endolyase) and exoeliminase (chondroitin sulfate ABC exolyase).
Hamai
A
A
Tokyo Research Institute, Seikagaku Corporation, Tateno 3-1253, Higashiyamato, Tokyo 207, Japan.
Hashimoto
N
N
Mochizuki
H
H
Kato
F
F
Makiguchi
Y
Y
Horie
K
K
Suzuki
S
S
D002808:Chondroitin Lyases / Q000302:isolation & purification / Q000378:metabolism*; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D003460:Crystallization; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D007527:Isoenzymes / Q000302:isolation & purification / Q000378:metabolism*; D009844:Oligosaccharides / Q000378:metabolism*; D011487:Protein Conformation; D011514:Proteus vulgaris / Q000201:enzymology*; D013379:Substrate Specificity; D014783:Viscosity
D007527:Isoenzymes; D009844:Oligosaccharides; D002808:Chondroitin Lyases