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Title: X-ray structure determination and deuteration of nattokinase.
Authors: Yanagisawa, Yasuhide
Chatake, Toshiyuki  kyouindb  KAKEN_id  orcid https://orcid.org/0000-0001-9208-9951 (unconfirmed)
Naito, Sawa
Ohsugi, Tadanori
Yatagai, Chieko
Sumi, Hiroyuki
Kawaguchi, Akio  kyouindb  KAKEN_id
Chiba-Kamosida, Kaori
Ogawa, Megumi
Adachi, Tatsumi
Morimoto, Yukio  kyouindb  KAKEN_id  orcid https://orcid.org/0000-0003-1919-6309 (unconfirmed)
Author's alias: 茶竹, 俊行
Keywords: nattokinase
Bacillus subtilis natto
Bacillus subtilis natto
X-ray structure
Issue Date: Nov-2013
Publisher: International Union of Crystallography
Journal title: Journal of synchrotron radiation
Volume: 20
Issue: Patt 6
Start page: 875
End page: 879
Abstract: Nattokinase (NK) is a strong fibrinolytic enzyme, which is produced in abundance by Bacillus subtilis natto. Although NK is a member of the subtilisin family, it displays different substrate specificity when compared with other subtilisins. The results of molecular simulations predict that hydrogen arrangements around Ser221 at the active site probably account for the substrate specificity of NK. Therefore, neutron crystallographic analysis should provide valuable information that reveals the enzymatic mechanism of NK. In this report, the X-ray structure of the non-hydrogen form of undeuterated NK was determined, and the preparation of deuterated NK was successfully achieved. The non-hydrogen NK structure was determined at 1.74 Å resolution. The three-dimensional structures of NK and subtilisin E from Bacillus subtilis DB104 are near identical. Deuteration of NK was carried out by cultivating Bacillus subtilis natto in deuterated medium. The D2O resistant strain of Bacillus subtilis natto was obtained by successive cultivation rounds, in which the concentration of D2O in the medium was gradually increased. NK was purified from the culture medium and its activity was confirmed by the fibrin plate method. The results lay the framework for neutron protein crystallography analysis.
Rights: © International Union of Crystallography
URI: http://hdl.handle.net/2433/179415
DOI(Published Version): 10.1107/S0909049513020700
PubMed ID: 24121331
Appears in Collections:Journal Articles

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