1ukq: Difference between revisions

Revision as of 16:25, 21 February 2008

Crystal structure of cyclodextrin glucanotransferase complexed with a pseudo-maltotetraose derived from acarbose

OverviewOverview

The stacking interaction between a tyrosine residue and the sugar ring at the catalytic subsite -1 is strictly conserved in the glycoside hydrolase family 13 enzymes. Replacing Tyr100 with leucine in cyclodextrin glycosyltransferase (CGTase) from Bacillus sp. 1011 to prevent stacking significantly decreased all CGTase activities. The adjacent stacking interaction with both Phe183 and Phe259 onto the sugar ring at subsite +2 is essentially conserved among CGTases. F183L/F259L mutant CGTase affects donor substrate binding and/or acceptor binding during transglycosylation [Nakamura et al. (1994) Biochemistry 33, 9929-9936]. To elucidate the precise role of carbohydrate/aromatic stacking interaction at subsites -1 and +2 on the substrate binding of CGTases, we analyzed the X-ray structures of wild-type (2.0 A resolution), and Y100L (2.2 A resolution) and F183L/F259L mutant (1.9 A resolution) CGTases complexed with the inhibitor, acarbose. The refined structures revealed that acarbose molecules bound to the Y100L mutant moved from the active center toward the side chain of Tyr195, and the hydrogen bonding and hydrophobic interaction between acarbose and subsites significantly diminished. The position of pseudo-tetrasaccharide binding in the F183L/F259L mutant was closer to the non-reducing end, and the torsion angles of glycosidic linkages at subsites -1 to +1 on molecule 1 and subsites -2 to -1 on molecule 2 significantly changed compared with that of each molecule of wild-type-acarbose complex to adopt the structural change of subsite +2. These structural and biochemical data suggest that substrate binding in the active site of CGTase is critically affected by the carbohydrate/aromatic stacking interaction with Tyr100 at the catalytic subsite -1 and that this effect is likely a result of cooperation between Tyr100 and Phe259 through stacking interaction with substrate at subsite +2.

About this StructureAbout this Structure

1UKQ is a Single protein structure of sequence from Bacillus sp. with as ligand. Active as Cyclomaltodextrin glucanotransferase, with EC number 2.4.1.19 Full crystallographic information is available from OCA.

ReferenceReference

Effects of essential carbohydrate/aromatic stacking interaction with Tyr100 and Phe259 on substrate binding of cyclodextrin glycosyltransferase from alkalophilic Bacillus sp. 1011., Haga K, Kanai R, Sakamoto O, Aoyagi M, Harata K, Yamane K, J Biochem. 2003 Dec;134(6):881-91. PMID:14769878

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-[[Image:1ukq.jpg|left|200px]]<br /><applet load="1ukq" size="450" color="white" frame="true" align="right" spinBox="true"
+[[Image:1ukq.jpg|left|200px]]<br /><applet load="1ukq" size="350" color="white" frame="true" align="right" spinBox="true"
 caption="1ukq, resolution 2.0&Aring;" />
 '''Crystal structure of cyclodextrin glucanotransferase complexed with a pseudo-maltotetraose derived from acarbose'''<br />
 ==Overview==
-The stacking interaction between a tyrosine residue and the sugar ring at, the catalytic subsite -1 is strictly conserved in the glycoside hydrolase, family 13 enzymes. Replacing Tyr100 with leucine in cyclodextrin, glycosyltransferase (CGTase) from Bacillus sp. 1011 to prevent stacking, significantly decreased all CGTase activities. The adjacent stacking, interaction with both Phe183 and Phe259 onto the sugar ring at subsite +2, is essentially conserved among CGTases. F183L/F259L mutant CGTase affects, donor substrate binding and/or acceptor binding during transglycosylation, [Nakamura et al. (1994) Biochemistry 33, 9929-9936]. To elucidate the, precise role of carbohydrate/aromatic stacking interaction at subsites -1, and +2 on the substrate binding of CGTases, we analyzed the X-ray, structures of wild-type (2.0 A resolution), and Y100L (2.2 A resolution), and F183L/F259L mutant (1.9 A resolution) CGTases complexed with the, inhibitor, acarbose. The refined structures revealed that acarbose, molecules bound to the Y100L mutant moved from the active center toward, the side chain of Tyr195, and the hydrogen bonding and hydrophobic, interaction between acarbose and subsites significantly diminished. The, position of pseudo-tetrasaccharide binding in the F183L/F259L mutant was, closer to the non-reducing end, and the torsion angles of glycosidic, linkages at subsites -1 to +1 on molecule 1 and subsites -2 to -1 on, molecule 2 significantly changed compared with that of each molecule of, wild-type-acarbose complex to adopt the structural change of subsite +2., These structural and biochemical data suggest that substrate binding in, the active site of CGTase is critically affected by the, carbohydrate/aromatic stacking interaction with Tyr100 at the catalytic, subsite -1 and that this effect is likely a result of cooperation between, Tyr100 and Phe259 through stacking interaction with substrate at subsite, +2.
+The stacking interaction between a tyrosine residue and the sugar ring at the catalytic subsite -1 is strictly conserved in the glycoside hydrolase family 13 enzymes. Replacing Tyr100 with leucine in cyclodextrin glycosyltransferase (CGTase) from Bacillus sp. 1011 to prevent stacking significantly decreased all CGTase activities. The adjacent stacking interaction with both Phe183 and Phe259 onto the sugar ring at subsite +2 is essentially conserved among CGTases. F183L/F259L mutant CGTase affects donor substrate binding and/or acceptor binding during transglycosylation [Nakamura et al. (1994) Biochemistry 33, 9929-9936]. To elucidate the precise role of carbohydrate/aromatic stacking interaction at subsites -1 and +2 on the substrate binding of CGTases, we analyzed the X-ray structures of wild-type (2.0 A resolution), and Y100L (2.2 A resolution) and F183L/F259L mutant (1.9 A resolution) CGTases complexed with the inhibitor, acarbose. The refined structures revealed that acarbose molecules bound to the Y100L mutant moved from the active center toward the side chain of Tyr195, and the hydrogen bonding and hydrophobic interaction between acarbose and subsites significantly diminished. The position of pseudo-tetrasaccharide binding in the F183L/F259L mutant was closer to the non-reducing end, and the torsion angles of glycosidic linkages at subsites -1 to +1 on molecule 1 and subsites -2 to -1 on molecule 2 significantly changed compared with that of each molecule of wild-type-acarbose complex to adopt the structural change of subsite +2. These structural and biochemical data suggest that substrate binding in the active site of CGTase is critically affected by the carbohydrate/aromatic stacking interaction with Tyr100 at the catalytic subsite -1 and that this effect is likely a result of cooperation between Tyr100 and Phe259 through stacking interaction with substrate at subsite +2.
 ==About this Structure==
-UKQ is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Bacillus_sp. Bacillus sp.] with CA as [http://en.wikipedia.org/wiki/ligand ligand]. Active as [http://en.wikipedia.org/wiki/Cyclomaltodextrin_glucanotransferase Cyclomaltodextrin glucanotransferase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=2.4.1.19 2.4.1.19] Full crystallographic information is available from [http://ispc.weizmann.ac.il/oca-bin/ocashort?id=1UKQ OCA].
+UKQ is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Bacillus_sp. Bacillus sp.] with <scene name='pdbligand=CA:'>CA</scene> as [http://en.wikipedia.org/wiki/ligand ligand]. Active as [http://en.wikipedia.org/wiki/Cyclomaltodextrin_glucanotransferase Cyclomaltodextrin glucanotransferase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=2.4.1.19 2.4.1.19] Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1UKQ OCA].
 ==Reference==
-Effects of essential carbohydrate/aromatic stacking interaction with Tyr100 and Phe259 on substrate binding of cyclodextrin glycosyltransferase from alkalophilic Bacillus sp. 1011., Haga K, Kanai R, Sakamoto O, Aoyagi M, Harata K, Yamane K, J Biochem (Tokyo). 2003 Dec;134(6):881-91. PMID:[http://ispc.weizmann.ac.il//pmbin/getpm?pmid=14769878 14769878]
+Effects of essential carbohydrate/aromatic stacking interaction with Tyr100 and Phe259 on substrate binding of cyclodextrin glycosyltransferase from alkalophilic Bacillus sp. 1011., Haga K, Kanai R, Sakamoto O, Aoyagi M, Harata K, Yamane K, J Biochem. 2003 Dec;134(6):881-91. PMID:[http://ispc.weizmann.ac.il//pmbin/getpm?pmid=14769878 14769878]
 [[Category: Bacillus sp.]]
 [[Category: Cyclomaltodextrin glucanotransferase]]
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 [[Category: cgtase]]
-''Page seeded by [http://ispc.weizmann.ac.il/oca OCA ] on Wed Nov 21 04:12:07 2007''
+''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Feb 21 15:25:37 2008''