Demonstration of catalytic proton acceptor of chitosanase from Paenibacillus fukuinensis by comprehensive analysis of mutant library |
| |
| Authors: | Danya Isogawa Takeshi Fukuda Kouichi Kuroda Hideo Kusaoke Hisashi Kimoto Shin-ichiro Suye and Mitsuyoshi Ueda |
| |
| Institution: | (1) Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Sakyo, Kyoto 606-8502, Japan;(2) Department of Applied Chemistry and Biotechnology, Graduate School of Engineering, University of Fukui, 3-9-1 Bunkyo, Fukui 910-8507, Japan;(3) Department of Environmental and Biotechnological Frontier Engineering, Fukui University of Technology, 3-6-1 Gakuen, Fukui 910-8505, Japan;(4) Department of Bioscience, Fukui Prefectural University, 4-1-1 Matsuokakenjojima, Eiheiji-cho, Yoshida-gun, Fukui 910-1195, Japan; |
| |
| Abstract: | Chitosanase from Paenibacillus fukuinensis D2 is an attractive enzyme, and it exhibits both chitosanase and β-1, 4 glucanase activities. In our previous study, we generated
P. fukuinensis chitosanase-displaying yeast cells using a yeast cell surface-displaying system. Chitosanase-displaying yeast can be utilized
as a chitosanase cluster without many time-consuming purification steps. In this study, using the system, we have investigated
whether Glu302, which is supposed as a putative proton acceptor, is an essential amino acid residue for exhibiting chitosanase
activity and analyzed the contribution of mutual interaction between Glu302 and Asn312 to the activity. A mutant library in
which Glu302 and Asn312 were comprehensively substituted by the other amino acid residues was constructed on the yeast cell
surface. From the results of chitosanase and β-1, 4 glucanase activity assays, we demonstrated that Glu302 was a proton acceptor
for chitosanase activity, and Asn312 also participated in the hydrolysis of chitosan and cellulose. |
| |
| Keywords: | |
| 本文献已被 SpringerLink 等数据库收录！ |
|
