1v3h
From Proteopedia
(New page: 200px<br /><applet load="1v3h" size="450" color="white" frame="true" align="right" spinBox="true" caption="1v3h, resolution 1.60Å" /> '''The roles of Glu186 ...) |
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| - | [[Image:1v3h.jpg|left|200px]]<br /><applet load="1v3h" size=" | + | [[Image:1v3h.jpg|left|200px]]<br /><applet load="1v3h" size="350" color="white" frame="true" align="right" spinBox="true" |
caption="1v3h, resolution 1.60Å" /> | caption="1v3h, resolution 1.60Å" /> | ||
'''The roles of Glu186 and Glu380 in the catalytic reaction of soybean beta-amylase'''<br /> | '''The roles of Glu186 and Glu380 in the catalytic reaction of soybean beta-amylase'''<br /> | ||
==Overview== | ==Overview== | ||
| - | It has previously been suggested that the glutamic acid residues Glu186 | + | It has previously been suggested that the glutamic acid residues Glu186 and Glu380 of soybean beta-amylase play critical roles as a general acid and a general base catalyst, respectively. In order to confirm the roles of Glu186 and Glu380, each residue was mutated to a glutamine residue and the crystal structures of the substrate (E186Q/maltopentaose) and product (E380Q/maltose) complexes were determined at resolutions of 1.6 Angstrom and 1.9 Angstrom, respectively. Both mutant enzymes exhibited 16,000- and 37,000-fold decreased activity relative to that of the wild-type enzyme. The crystal structure of the E186Q/maltopentaose complex revealed an unambiguous five-glucose unit at subsites -2 to +3. Two maltose molecules bind on subsites -2 to -1 and +2 to +3 in the E380Q/maltose complex, whereas they bind in tandem to -2 to -1 and +1 to +2 in the wild-type/maltose complex. The conformation of the glucose residue at subsite -1 was identified as a stable (4)C(1) alpha-anomer in the E380Q/maltose complex, whereas a distorted ring conformation was observed in the wild-type/maltose complex. The side-chain movement of Gln380 to the position of a putative attacking water molecule seen in the wild-type enzyme caused the inactivation of the E380Q mutant and an altered binding pattern of maltose molecules. These results confirm the critical roles played by Glu186 in the donation of a proton to the glycosidic oxygen of the substrate, and by Glu380 in the activation of an attacking water molecule. The observed difference between the backbones of E186Q/maltopentaose and E380Q/maltose in terms of Thr342 suggests that the side-chain of Thr342 may stabilize the deprotonated form of Glu186 after the cleavage of the glycosidic bond. |
==About this Structure== | ==About this Structure== | ||
| - | 1V3H is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Glycine_max Glycine max] with SO4 as [http://en.wikipedia.org/wiki/ligand ligand]. Active as [http://en.wikipedia.org/wiki/Beta-amylase Beta-amylase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=3.2.1.2 3.2.1.2] Full crystallographic information is available from [http:// | + | 1V3H is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Glycine_max Glycine max] with <scene name='pdbligand=SO4:'>SO4</scene> as [http://en.wikipedia.org/wiki/ligand ligand]. Active as [http://en.wikipedia.org/wiki/Beta-amylase Beta-amylase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=3.2.1.2 3.2.1.2] Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1V3H OCA]. |
==Reference== | ==Reference== | ||
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[[Category: Single protein]] | [[Category: Single protein]] | ||
[[Category: Adachi, M.]] | [[Category: Adachi, M.]] | ||
| - | [[Category: Kang, Y | + | [[Category: Kang, Y N.]] |
[[Category: Mikami, B.]] | [[Category: Mikami, B.]] | ||
[[Category: Utsumi, S.]] | [[Category: Utsumi, S.]] | ||
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[[Category: (beta/alpha)8 barrel]] | [[Category: (beta/alpha)8 barrel]] | ||
| - | ''Page seeded by [http:// | + | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Feb 21 15:31:08 2008'' |
Revision as of 13:31, 21 February 2008
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The roles of Glu186 and Glu380 in the catalytic reaction of soybean beta-amylase
Overview
It has previously been suggested that the glutamic acid residues Glu186 and Glu380 of soybean beta-amylase play critical roles as a general acid and a general base catalyst, respectively. In order to confirm the roles of Glu186 and Glu380, each residue was mutated to a glutamine residue and the crystal structures of the substrate (E186Q/maltopentaose) and product (E380Q/maltose) complexes were determined at resolutions of 1.6 Angstrom and 1.9 Angstrom, respectively. Both mutant enzymes exhibited 16,000- and 37,000-fold decreased activity relative to that of the wild-type enzyme. The crystal structure of the E186Q/maltopentaose complex revealed an unambiguous five-glucose unit at subsites -2 to +3. Two maltose molecules bind on subsites -2 to -1 and +2 to +3 in the E380Q/maltose complex, whereas they bind in tandem to -2 to -1 and +1 to +2 in the wild-type/maltose complex. The conformation of the glucose residue at subsite -1 was identified as a stable (4)C(1) alpha-anomer in the E380Q/maltose complex, whereas a distorted ring conformation was observed in the wild-type/maltose complex. The side-chain movement of Gln380 to the position of a putative attacking water molecule seen in the wild-type enzyme caused the inactivation of the E380Q mutant and an altered binding pattern of maltose molecules. These results confirm the critical roles played by Glu186 in the donation of a proton to the glycosidic oxygen of the substrate, and by Glu380 in the activation of an attacking water molecule. The observed difference between the backbones of E186Q/maltopentaose and E380Q/maltose in terms of Thr342 suggests that the side-chain of Thr342 may stabilize the deprotonated form of Glu186 after the cleavage of the glycosidic bond.
About this Structure
1V3H is a Single protein structure of sequence from Glycine max with as ligand. Active as Beta-amylase, with EC number 3.2.1.2 Full crystallographic information is available from OCA.
Reference
The roles of Glu186 and Glu380 in the catalytic reaction of soybean beta-amylase., Kang YN, Adachi M, Utsumi S, Mikami B, J Mol Biol. 2004 Jun 18;339(5):1129-40. PMID:15178253
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