Purification and properties of a xylanase from Cellvibrio gilvus that hydrolyzes p-nitrophenyl cellooligosaccharides.
An enzyme component that hydrolyzes pNP-G2 but not CMC has been isolated from a culture broth of Cellvibrio gilvus by a multi-step procedure involving Butyl-Toyopearl, DEAE-Toyopearl, and CM-Toyopearl chromatographies. The purified enzyme gave a single protein band on native, SDS-, and IEF-PAGE. The enzyme had a molecular weight of 40,000, an isoelectric point of 5.0, an optimum pH of 6.5, and an optimum temperature of 55 degrees C. It was stable from pH 4.0 to 9.0 at 37 degrees C for 1 hr and below 50 degrees C for 30 min. It hydrolyzed agluconic bonds not only of pNP-G2 but also of pNP-G3, pNP-G4, and pNP-G5. Cellooligosaccharides with D.P. of 3 to 5 were not hydrolyzed at all. Instead, the enzyme hydrolyzed xylan 4 times as fast as pNP-G2. Both HgCl2 and p-chloromercuribenzoic acid inhibited the two activities completely. Xylan inhibited the hydrolysis of pNP-G2 competitively. From these results, the purified enzyme was considered to be a unique xylanase that hydrolyzed the agluconic bonds of pNP-Gn.
Haga
K
K
National Food Research Institute, Ibaraki, Japan.
Kitaoka
M
M
Kashiwagi
Y
Y
Sasaki
T
T
Taniguchi
H
H
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000037:antagonists & inhibitors / Q000302:isolation & purification* / Q000378:metabolism; D002850:Chromatography, Gel / Q000379:methods; D005960:Glucosides / Q000378:metabolism; D006026:Glycoside Hydrolases / Q000037:antagonists & inhibitors / Q000302:isolation & purification* / Q000378:metabolism; D006088:Gram-Negative Aerobic Bacteria / Q000201:enzymology*; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000378:metabolism*; D013696:Temperature; D043325:Xylan Endo-1,3-beta-Xylosidase; D014990:Xylans / Q000494:pharmacology
D001426:Bacterial Proteins; D005960:Glucosides; D009844:Oligosaccharides; D014990:Xylans; C034170:4-nitrophenyl beta-cellobioside; D006026:Glycoside Hydrolases; D043325:Xylan Endo-1,3-beta-Xylosidase