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Detection of major xylanase-containing cellulose-binding domain from Penicillium verruculosum by combination of chromatofocusing and limited proteolysisстатья
Информация о цитировании статьи получена из
Web of Science,
Scopus
Статья опубликована в журнале из списка Web of Science и/или Scopus
Дата последнего поиска статьи во внешних источниках: 18 июля 2013 г.
- Авторы: Berlin A.G., Gusakov A.V., Sinitsyna O.A., Sinitsyn A.P.
- Журнал: Applied Biochemistry and Biotechnology
- Том: 88
- Номер: 1-3
- Год издания: 2000
- Издательство: Humana Press, Inc.
- Местоположение издательства: United States
- Первая страница: 345
- Последняя страница: 352
- DOI: 10.1385/ABAB:88:1-3:345
- Аннотация: Adsorption on microcrystalline cellulose of enzyme components of cellulase complex from Penicillium verruculosum was studied by chromatofocusing on a Mono P column. The most strongly adsorbed and major component was identified as xylanase (XYN) with MW 65 kDa and pi 4.5. The high adsorption degree of XYN on cellulose indicated the possible presence of a cellulose-binding domain in the molecular structure. Limited proteolysis of XYN with papain was carried out. Kinetics of proteolysis was monitored by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis and measuring activities toward insoluble xylan and 4-methyl-umbelliferyl-beta-D-lactoside (MUF-LAC). During the proteolysis, formation of two polypeptides with MW 51 and 14 kDa was observed. No loss of activity toward the soluble substrate was observed, whereas the activity toward xylan decreased rapidly. Adsorption distribution coefficient (K-d) of the core protein separated by gel-filtration was found to be 15 times lower than the K-d, for the initial nondigested XYN (0.02 and 0.29 L/g, respectively). The activity of core protein toward insoluble xylan was close to zero, whereas the activity toward MUF-LAC was close to that exhibited by the original enzyme. The results presented indicate a bifunctional organization of XYN, where one domain acts as a binding anchor for insoluble substrates and the other, localized in the core protein, contains the active site.
- Добавил в систему: Гусаков Александр Васильевич