TAILIEUCHUNG - Báo cáo khoa học: Crystal structure of a family 16 endoglucanase from the hyperthermophile Pyrococcus furiosus – structural basis of substrate recognition

Bacterial and archaeal endo-b-1,3-glucanases that belong to glycoside hydrolase family 16 share a b-jelly-roll fold, but differ significantly in sequence and in substrate specificity. The crystal structure of the laminarin-ase (EC ) from the hyperthermophilic archaeonPyrococcus furiosus (pfLamA) has been determined at A˚ resolution by molecular replace-ment. ThepfLamA structure reveals a kink of six residues (72–77) at the entrance of the catalytic cleft. | Crystal structure of a family 16 endoglucanase from the hyperthermophile Pyrococcus furiosus - structural basis of substrate recognition 2 2 2 Andrea Ilari Annarita Fiorillo Sebastiana Angelaccio Rita Florio Roberta Chiaraluce John van der Oost4 and Valerio Consalvi2 1 CNR Institute of Molecular Biology and Pathology Italy 2 Dipartimento di Scienze Biochimiche A. Rossi Fanelli Sapienza Universita di Roma Italy 3 Dipartimento di Scienze e Tecnologie Biomediche University degli Studi Dell Aquila 4 Laboratory of Microbiology Wageningen University The Netherlands Keywords b-1 3 glucan specificity crystal structure endoglucanase laminarin lichenan Correspondence A. Ilari Institute of Molecular Biology and Pathology CNR c o Dipartimento di Scienze Biochimiche A. Rossi Fanelli Sapienza University di Roma Piazzale Aldo Moro 5 00185 Rome Italy Fax 39064440062 Tel 390649910910 E-mail These authors contributed equally to this work Bacterial and archaeal endo-b-1 3-glucanases that belong to glycoside hydrolase family 16 share a b-jelly-roll fold but differ significantly in sequence and in substrate specificity. The crystal structure of the laminarin-ase EC from the hyperthermophilic archaeon Pyrococcus furiosus pfLamA has been determined at A resolution by molecular replacement. The pfLamA structure reveals a kink of six residues 72-77 at the entrance of the catalytic cleft. This peptide is absent in the endoglucanases from alkaliphilic Nocardiopsis sp. strain F96 and Bacillus macerans two proteins displaying an overall fold similar to that of pfLamA but with different substrate specificity. A deletion mutant of pfLamA lacking residues 72-75 hydrolyses the mixed-linkage b-1 3-1 4-glucan lichenan 10 times more efficiently than the wild-type protein indicating the importance of the kink in substrate preference. Received 27 October 2008 revised 5 December 2008 accepted 9 December 2008 doi .

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