Purification, properties, and transglycosylation reaction of .BETA.-N-acetylhexosaminidase from Nocardia orientalis.

Abstract

/J-/V-Acetylhexosaminidase (EC 3.2.1.52) from the culture filtrate of Nocardia orientalis was purified to homogeneity by precipitation with ammonium sulfate followed by column chromatography on CM-Sephadex, Bio-Gel P-60, and phenyl-Sepharose CL-4B. The molecular weight of the enzyme was about 56,000 by gel filtration and 54,000 by SDSpolyacrylamide gel electrophoresis. The enzyme showed about 1.6-fold higher j?-/V-acetylglucosaminidase activity than /?-/V-acetyIgaIactosaminidase activity. The optimum pH and temperature were 5.0 and around 70-75 C for PNP-GlcNAc, and 4.0 and 60 C for PNP-GalNAc. The enzyme was stable in the pH range from 4.0 to 8.0 and below 45 C.The enzyme hydrolyzed /V-acetyl-chitooligosaccharides, di-7V-acetyl-chitobiose through hexa-/Vacetyl-chitohexaose. The enzyme showed glycosyl transferase activity during the hydrolysis of di-/Vacetyl-chitobiose.Twomajor transfer products were isolated and identified as the /Hl-»6)-linked disaccharide of N-acetylglucosamine and tri-/V-acetyl-chitotriose.jS-iV-Acetylhexosaminidase (EC 3.2.

Purification and characterization of ββ‐N‐acetylhexosaminidase from the phytopathogenic fungusBipolaris sorokiniana

“…The enzyme b-N-acetylhexosaminidase was purified from the fungus B. sorokiniana. Purification was about 70-fold, yielding 41%, similar to the data described for other microorganisms (Eriquez and Pisano 1979;Jones and Kosman 1980;Nanjo et al 1990). The purification step of gel filtration was effective where the major quantity of protein eluted after the enzyme.…”

“…The enzyme β‐N‐acetylhexosaminidase was purified from the fungus B. sorokiniana . Purification was about 70‐fold, yielding 41%, similar to the data described for other micro‐organisms (Eriquez & Pisano 1979; Jones & Kosman 1980; Nanjo et al . 1990).…”

Chitinolytic enzymes: An appraisal as a product of commercial potential

“…A similar type of transglycosylation was also reported for a chitinase from T. reesei and chitinase from Streptomyces kurssanovii . The purified N‐ acetylglucosaminidase of N. orientalis and β‐ N ‐acetylhexosaminidase of Penicillium oxalicum were found to possess transglycosylation activity in addition to the hydrolytic activity . The latter enzyme effectively transferred GlcNAc to various alcohols in the presence of N ‐acetylchitooligomers as donors.…”

Purification and characterization of a chitosanase from Streptomyces N174

“…For each enzyme studied, a strong relationship between the degree of N-acetylation of the substrate and the relative enzyme activity was observed. Whereas, chitinase and lysozyme preferentially attack highly N-acetylated polymers, chitosanases hydrolyse more efficiently chitosans with a low N-acetyl content (Fenton and Eveleigh 1981;Ohtakara et al 1988;Sashiwa et al 1990;Nanjo et al 1990a;b). In partially N-acetylated chitosans, chitinases specifically cleave the N-acetyl-l~-D-glucosaminidic bonds Mitsutomi et al 1990).…”