Purification of sulfated fucoglucuronomannan lyase from bacterial strain of Fucobacter marina and study of appropriate conditions for its enzyme digestion.
A marine bacterial strain, Fucobacter marina, produced extracellular sulfated fucoglucuronomannan (SFGM) lyase when cultivated in the presence of crude SFGM obtained from fucoidan of Kjellmaniella crassifolia (brown algae) by cetyl pyridinium chloride fractionation. For the SFGM lyase assay, SFGM fraction separated from K. crassifolia fucoidan by anion exchange column chromatography was used as the substrate. The extracellular SFGM lyase was purified to homogeneity on an electrophoresis gel with 4240-fold purity at 13.8% yield. The enzyme proved to be a monomer, since gel filtration and sodium dodecyl sulfate polyacrylamide gel electrophoresis gave the same relative molecular mass of 67,000. The enzyme specifically digested SFGM but did not digest any other uronic-acid-containing polysaccharides tested. The optimum conditions for the enzyme reaction were around pH 7.5, 43 degrees C, and 0.4 M NaCl concentration. The enzyme was strongly inhibited by CuCl(2) and ZnCl(2), and also by some sulfhydryl reagents.
Sakai
Takeshi
T
Takara Bio Inc., Biotechnology Research Laboratories, Zaifu-cho 82-4, Hirosaki-shi, Aomori-ken, 036-8216, Japan. sakait@takara-bio.co.jp
Kimura
Hitomi
H
Kato
Ikunoshin
I
D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D041965:Flavobacteriaceae / Q000201:enzymology*; D005643:Fucose / Q000031:analogs & derivatives* / Q000302:isolation & purification; D006863:Hydrogen-Ion Concentration; D008361:Mannosidases / Q000302:isolation & purification* / Q000378:metabolism; D000459:Phaeophyceae / Q000737:chemistry; D011134:Polysaccharides / Q000737:chemistry; D013379:Substrate Specificity
D011134:Polysaccharides; D005643:Fucose; C007789:fucoidan; D008361:Mannosidases; C051635:endo-alpha-D-mannosidase