title: Purification and characterization of a novel beta-agarase from an alkalophilic bacterium, Alteromonas sp. E-1.
abstract: A novel beta-agarase (EC 3.2.1.81) was purified from an agar-degrading alkalophilic bacterium, Alteromonas sp. E-1 isolated from the soil. This enzyme was obtained from a cell-free extract after sonication and purified 40.9-fold through treatment with streptomycin, ammonium sulfate fractionation and successive chromatography on anion-exchange and gel filtration columns. The molecular weight was estimated to be 82 kDa by SDS-polyacrylamide gel electrophoresis and 180 kDa by Superdex 200 gel filtration. The enzyme was inhibited by Mn2+, Cu2+, Fe2+, Zn2+ and Hg2+, and activated by K+, Na+ and EDTA, and its optimum pH and temperature for agarose degradation were 7.5 and 40 degrees C, respectively. This beta-agarase hydrolyzed agarose with rapid reduction of viscosity, and neoagarobiose [O-3,6-anhydro-alpha-L-galactopyranosyl(1-->3)-D-galactose] was detected from the early stage of the reaction. Neoagarobiose as the final product was selectively released from agarose, neoagarohexaose and neoagarotetraose by the reaction with this beta-agarase. This observation was different from that of other beta-agarases which produced mixtures of neoagarobiose and neoagarotetraose as the final hydrolysis products. The N-terminal amino acid sequence of this beta-agarase shows no homology to those of other beta-agarases that were so far reported.
authors.affiliation: Department of Applied Chemistry, School of Science and Engineering, Waseda University, 3-4-1 Ohkubo, Tokyo 169-8555, Japan.
authors.forename: K
authors.initials: K
authors.lastname: Kirimura
authors.forename: N
authors.initials: N
authors.lastname: Masuda
authors.forename: Y
authors.initials: Y
authors.lastname: Iwasaki
authors.forename: H
authors.initials: H
authors.lastname: Nakagawa
authors.forename: R
authors.initials: R
authors.lastname: Kobayashi
authors.forename: S
authors.initials: S
authors.lastname: Usami