1gbt
From Proteopedia
(New page: 200px<br /><applet load="1gbt" size="450" color="white" frame="true" align="right" spinBox="true" caption="1gbt, resolution 2.0Å" /> '''STRUCTURE OF AN ACYL-...) |
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| - | [[Image:1gbt.jpg|left|200px]]<br /><applet load="1gbt" size=" | + | [[Image:1gbt.jpg|left|200px]]<br /><applet load="1gbt" size="350" color="white" frame="true" align="right" spinBox="true" |
caption="1gbt, resolution 2.0Å" /> | caption="1gbt, resolution 2.0Å" /> | ||
'''STRUCTURE OF AN ACYL-ENZYME INTERMEDIATE DURING CATALYSIS: (GUANIDINOBENZOYL) TRYPSIN'''<br /> | '''STRUCTURE OF AN ACYL-ENZYME INTERMEDIATE DURING CATALYSIS: (GUANIDINOBENZOYL) TRYPSIN'''<br /> | ||
==Overview== | ==Overview== | ||
| - | The crystal and molecular structure of trypsin at a transiently stable | + | The crystal and molecular structure of trypsin at a transiently stable intermediate step during catalysis has been determined by X-ray diffraction methods. Bovine trypsin cleaved the substrate p-nitrophenyl p-guanidinobenzoate during crystallization under conditions in which the acyl-enzyme intermediate, (guanidinobenzoyl)trypsin, was stable. Orthorhombic crystals formed in space group P2(1)2(1)2(1), with a = 63.74, b = 63.54, and c = 68.93 A. This is a crystal form of bovine trypsin for which a molecular structure has not been reported. Diffraction data were measured with a FAST (Enraf Nonius) diffractometer. The structure was refined to a crystallographic residual of R = 0.16 for data in the resolution range 7.0-2.0 A. The refined model of (guanidinobenzoyl)trypsin provides insight into the structural basis for its slow rate of deacylation, which in solution at 25 degrees C and pH 7.4 exhibits a t1/2 of 12 h. In addition to the rotation of the Ser-195 hydroxyl away from His-157, C beta of Ser-195 moves 0.7 A toward Asp-189 at the bottom of the active site, with respect to the native structure. This allows formation of energetically favorable H bonds and an ion pair between the carboxylate of Asp-189 and the guanidino group of the substrate. This movement is dictated by the rigidity of the aromatic ring in guanidinobenzoate--model-building indicates that this should not occur when arginine, with its more flexible aliphatic backbone, forms the ester bond with Ser-195. As a consequence, highly ordered water molecules in the active site are no longer close enough to the scissile ester bond to serve as potential nucleophiles for hydrolysis.(ABSTRACT TRUNCATED AT 250 WORDS) |
==About this Structure== | ==About this Structure== | ||
| - | 1GBT is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Bos_taurus Bos taurus] with CA, SO4 and GBS as [http://en.wikipedia.org/wiki/ligands ligands]. Active as [http://en.wikipedia.org/wiki/Trypsin Trypsin], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=3.4.21.4 3.4.21.4] Full crystallographic information is available from [http:// | + | 1GBT is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Bos_taurus Bos taurus] with <scene name='pdbligand=CA:'>CA</scene>, <scene name='pdbligand=SO4:'>SO4</scene> and <scene name='pdbligand=GBS:'>GBS</scene> as [http://en.wikipedia.org/wiki/ligands ligands]. Active as [http://en.wikipedia.org/wiki/Trypsin Trypsin], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=3.4.21.4 3.4.21.4] Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1GBT OCA]. |
==Reference== | ==Reference== | ||
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[[Category: Single protein]] | [[Category: Single protein]] | ||
[[Category: Trypsin]] | [[Category: Trypsin]] | ||
| - | [[Category: Singer, P | + | [[Category: Singer, P T.]] |
| - | [[Category: Sweet, R | + | [[Category: Sweet, R M.]] |
[[Category: CA]] | [[Category: CA]] | ||
[[Category: GBS]] | [[Category: GBS]] | ||
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[[Category: hydrolase(serine proteinase)]] | [[Category: hydrolase(serine proteinase)]] | ||
| - | ''Page seeded by [http:// | + | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Feb 21 12:48:24 2008'' |
Revision as of 10:48, 21 February 2008
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STRUCTURE OF AN ACYL-ENZYME INTERMEDIATE DURING CATALYSIS: (GUANIDINOBENZOYL) TRYPSIN
Overview
The crystal and molecular structure of trypsin at a transiently stable intermediate step during catalysis has been determined by X-ray diffraction methods. Bovine trypsin cleaved the substrate p-nitrophenyl p-guanidinobenzoate during crystallization under conditions in which the acyl-enzyme intermediate, (guanidinobenzoyl)trypsin, was stable. Orthorhombic crystals formed in space group P2(1)2(1)2(1), with a = 63.74, b = 63.54, and c = 68.93 A. This is a crystal form of bovine trypsin for which a molecular structure has not been reported. Diffraction data were measured with a FAST (Enraf Nonius) diffractometer. The structure was refined to a crystallographic residual of R = 0.16 for data in the resolution range 7.0-2.0 A. The refined model of (guanidinobenzoyl)trypsin provides insight into the structural basis for its slow rate of deacylation, which in solution at 25 degrees C and pH 7.4 exhibits a t1/2 of 12 h. In addition to the rotation of the Ser-195 hydroxyl away from His-157, C beta of Ser-195 moves 0.7 A toward Asp-189 at the bottom of the active site, with respect to the native structure. This allows formation of energetically favorable H bonds and an ion pair between the carboxylate of Asp-189 and the guanidino group of the substrate. This movement is dictated by the rigidity of the aromatic ring in guanidinobenzoate--model-building indicates that this should not occur when arginine, with its more flexible aliphatic backbone, forms the ester bond with Ser-195. As a consequence, highly ordered water molecules in the active site are no longer close enough to the scissile ester bond to serve as potential nucleophiles for hydrolysis.(ABSTRACT TRUNCATED AT 250 WORDS)
About this Structure
1GBT is a Single protein structure of sequence from Bos taurus with , and as ligands. Active as Trypsin, with EC number 3.4.21.4 Full crystallographic information is available from OCA.
Reference
Structure of an acyl-enzyme intermediate during catalysis: (guanidinobenzoyl)trypsin., Mangel WF, Singer PT, Cyr DM, Umland TC, Toledo DL, Stroud RM, Pflugrath JW, Sweet RM, Biochemistry. 1990 Sep 11;29(36):8351-7. PMID:2252895
Page seeded by OCA on Thu Feb 21 12:48:24 2008
Categories: Bos taurus | Single protein | Trypsin | Singer, P T. | Sweet, R M. | CA | GBS | SO4 | Hydrolase(serine proteinase)
