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Hokkaido University Collection of Scholarly and Academic Papers >
Graduate School of Agriculture / Faculty of Agriculture >
Peer-reviewed Journal Articles, etc >
The N-terminal region of the starch-branching enzyme from Phaseolus vulgaris L. is essential for optimal catalysis and structural stability
| Title: | The N-terminal region of the starch-branching enzyme from Phaseolus vulgaris L. is essential for optimal catalysis and structural stability |
| Authors: | Hamada, Shigeki Browse this author | | Ito, Hiroyuki Browse this author | | Ueno, Hiroshi Browse this author | | Takeda, Yasuhito Browse this author | | Matsui, Hirokazu Browse this author →KAKEN DB |
| Keywords: | Kidney bean | | Phaseolus vulgaris L. | | Leguminosae | | Chimeric enzyme | | Site-directed mutagenesis | | Starch-branching enzyme | | Kinetics |
| Issue Date: | May-2007 |
| Publisher: | Elsevier Ltd. |
| Journal Title: | Phytochemistry |
| Volume: | 68 |
| Issue: | 10 |
| Start Page: | 1367 |
| End Page: | 1375 |
| Publisher DOI: | 10.1016/j.phytochem.2007.02.024 |
| PMID: | 17408708 |
| Abstract: | Starch-branching enzymes (SBEs) play a pivotal role in determining the fine structure of starch by catalyzing the syntheses of α-1,6-branch points. They are the members of the α-amylase family and have four conserved regions in a central (β/α)8 barrel, including the catalytic sites. Although the role of the catalytic barrel domain of an SBE is known, that of its N- and C-terminal regions remain unclear. We have previously shown that the C-terminal regions of the two SBE isozymes (designated as PvSBE1 and PvSBE2) from kidney bean (Phaseolus vulgaris L.) have different roles in branching enzyme activity. To understand the contribution of the N-terminal region to catalysis, six chimeric enzymes were constructed between PvSBE1 and PvSBE2. Only one enzyme (1Na/2Nb)-II, in which a portion of the N-terminal region of PvSBE2 was substituted by the corresponding region of PvSBE1, retained 6% of the PvSBE2 activity. The N-terminal truncated form (ΔN46-PvSBE2), lacking 46 N-terminal residues of PvSBE2, lost enzyme activity and stability to proteolysis. To investigate the possible function of this region, three residues (Asp-15, His-24, and Arg-28) among these 46 residues were subjected to site-directed mutagenesis. The purified mutant enzymes showed nearly the same Km values as PvSBE2 but had lower Vmax values and heat stabilities than PvSBE2. These results suggest that the N-terminal region of the kidney bean SBE is essential for maximum enzyme activity and thermostability. |
| Relation: | http://www.sciencedirect.com/science/journal/00319422 |
| Type: | article (author version) |
| URI: | http://hdl.handle.net/2115/27965 |
| Appears in Collections: | 農学院・農学研究院 (Graduate School of Agriculture / Faculty of Agriculture) > 雑誌発表論文等 (Peer-reviewed Journal Articles, etc)
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Submitter: 伊藤 浩之
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