Thermophilic and halophilic β-agarase from a halophilic archaeon Halococcus sp. 197A.
An agar-degrading archaeon Halococcus sp. 197A was isolated from a solar salt sample. The agarase was purified by hydrophobic column chromatography using a column of TOYOPEARL Phenyl-650 M. The molecular mass of the purified enzyme, designated as Aga-HC, was ~55 kDa on both SDS-PAGE and gel-filtration chromatography. Aga-HC released degradation products in the order of neoagarohexose, neoagarotetraose and small quantity of neoagarobiose, indicating that Aga-HC was a β-type agarase. Aga-HC showed a salt requirement for both stability and activity, being active from 0.3 M NaCl, with maximal activity at 3.5 M NaCl. KCl supported similar activities as NaCl up to 3.5 M, and LiCl up to 2.5 M. These monovalent salts could not be substituted by 3.5 M divalent cations, CaCl2 or MgCl2. The optimal pH was 6.0. Aga-HC was thermophilic, with optimum temperature of 70 °C. Aga-HC retained approximately 90 % of the initial activity after incubation for 1 hour at 65-80 °C, and retained 50 % activity after 1 hour at 95 °C. In the presence of additional 10 mM CaCl2, approximately 17 % remaining activity was detected after 30 min at 100 °C. This is the first report on agarase purified from Archaea.
Bio-Nano Electronics Research Center, Toyo University, 2100 Kujirai, Kawagoe, Saitama, 350-8585, Japan, minehiro@toyo.jp.
Hiroaki
H
Minegishi
Yasuhiro
Y
Shimane
Akinobu
A
Echigo
Yukari
Y
Ohta
Yuji
Y
Hatada
Masahiro
M
Kamekura
Tadashi
T
Maruyama
Ron
R
Usami
D019843:Archaeal Proteins / Q000737:chemistry / Q000378:metabolism*; D004795:Enzyme Stability; D006026:Glycoside Hydrolases / Q000737:chemistry / Q000378:metabolism*; D019625:Halococcus / Q000201:enzymology*; D013379:Substrate Specificity
D019843:Archaeal Proteins; D006026:Glycoside Hydrolases; C019624:agarase