1yjs
From Proteopedia
(New page: 200px<br /><applet load="1yjs" size="450" color="white" frame="true" align="right" spinBox="true" caption="1yjs, resolution 2.00Å" /> '''K226Q Mutant Of Seri...) |
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| - | [[Image:1yjs.gif|left|200px]]<br /><applet load="1yjs" size=" | + | [[Image:1yjs.gif|left|200px]]<br /><applet load="1yjs" size="350" color="white" frame="true" align="right" spinBox="true" |
caption="1yjs, resolution 2.00Å" /> | caption="1yjs, resolution 2.00Å" /> | ||
'''K226Q Mutant Of Serine Hydroxymethyltransferase From B. Stearothermophilus, Complex With Glycine'''<br /> | '''K226Q Mutant Of Serine Hydroxymethyltransferase From B. Stearothermophilus, Complex With Glycine'''<br /> | ||
==Overview== | ==Overview== | ||
| - | Serine hydroxymethyltransferase (SHMT), a pyridoxal 5'-phosphate | + | Serine hydroxymethyltransferase (SHMT), a pyridoxal 5'-phosphate (PLP)-dependent enzyme catalyzes the reversible conversion of l-Ser and tetrahydropteroylglutamate (H(4)PteGlu) to Gly and 5,10-methylene tetrahydropteroylglutamate (CH(2)-H(4)PteGlu). Biochemical and structural studies on this enzyme have implicated several residues in the catalytic mechanism, one of them being the active site lysine, which anchors PLP. It has been proposed that this residue is crucial for product expulsion. However, in other PLP-dependent enzymes, the corresponding residue has been implicated in the proton abstraction step of catalysis. In the present investigation, Lys-226 of Bacillus stearothermophilus SHMT (bsSHMT) was mutated to Met and Gln to evaluate the role of this residue in catalysis. The mutant enzymes contained 1 mol of PLP per mol of subunit suggesting that Schiff base formation with lysine is not essential for PLP binding. The 3D structure of the mutant enzymes revealed that PLP was bound at the active site in an orientation different from that of the wild-type enzyme. In the presence of substrate, the PLP ring was in an orientation superimposable with that of the external aldimine complex of wild-type enzyme. However, the mutant enzymes were inactive, and the kinetic analysis of the different steps of catalysis revealed that there was a drastic reduction in the rate of formation of the quinonoid intermediate. Analysis of these results along with the crystal structures suggested that K-226 is responsible for flipping of PLP from one orientation to another which is crucial for H(4)PteGlu-dependent Calpha-Cbeta bond cleavage of l-Ser. |
==About this Structure== | ==About this Structure== | ||
| - | 1YJS is a [http://en.wikipedia.org/wiki/Protein_complex Protein complex] structure of sequences from [http://en.wikipedia.org/wiki/Geobacillus_stearothermophilus Geobacillus stearothermophilus] with PLP as [http://en.wikipedia.org/wiki/ligand ligand]. Active as [http://en.wikipedia.org/wiki/Glycine_hydroxymethyltransferase Glycine hydroxymethyltransferase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=2.1.2.1 2.1.2.1] Full crystallographic information is available from [http:// | + | 1YJS is a [http://en.wikipedia.org/wiki/Protein_complex Protein complex] structure of sequences from [http://en.wikipedia.org/wiki/Geobacillus_stearothermophilus Geobacillus stearothermophilus] with <scene name='pdbligand=PLP:'>PLP</scene> as [http://en.wikipedia.org/wiki/ligand ligand]. Active as [http://en.wikipedia.org/wiki/Glycine_hydroxymethyltransferase Glycine hydroxymethyltransferase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=2.1.2.1 2.1.2.1] Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1YJS OCA]. |
==Reference== | ==Reference== | ||
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[[Category: Glycine hydroxymethyltransferase]] | [[Category: Glycine hydroxymethyltransferase]] | ||
[[Category: Protein complex]] | [[Category: Protein complex]] | ||
| - | [[Category: Appaji, R | + | [[Category: Appaji, R N.]] |
[[Category: Bhavani, S.]] | [[Category: Bhavani, S.]] | ||
| - | [[Category: Jala, V | + | [[Category: Jala, V R.]] |
[[Category: Kaul, P.]] | [[Category: Kaul, P.]] | ||
[[Category: Prakash, V.]] | [[Category: Prakash, V.]] | ||
[[Category: Purnima, K.]] | [[Category: Purnima, K.]] | ||
| - | [[Category: Savithri, H | + | [[Category: Savithri, H S.]] |
| - | [[Category: Subramanya, H | + | [[Category: Subramanya, H S.]] |
[[Category: Trivedi, V.]] | [[Category: Trivedi, V.]] | ||
[[Category: PLP]] | [[Category: PLP]] | ||
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[[Category: shmt]] | [[Category: shmt]] | ||
| - | ''Page seeded by [http:// | + | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Feb 21 16:06:03 2008'' |
Revision as of 14:06, 21 February 2008
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K226Q Mutant Of Serine Hydroxymethyltransferase From B. Stearothermophilus, Complex With Glycine
Overview
Serine hydroxymethyltransferase (SHMT), a pyridoxal 5'-phosphate (PLP)-dependent enzyme catalyzes the reversible conversion of l-Ser and tetrahydropteroylglutamate (H(4)PteGlu) to Gly and 5,10-methylene tetrahydropteroylglutamate (CH(2)-H(4)PteGlu). Biochemical and structural studies on this enzyme have implicated several residues in the catalytic mechanism, one of them being the active site lysine, which anchors PLP. It has been proposed that this residue is crucial for product expulsion. However, in other PLP-dependent enzymes, the corresponding residue has been implicated in the proton abstraction step of catalysis. In the present investigation, Lys-226 of Bacillus stearothermophilus SHMT (bsSHMT) was mutated to Met and Gln to evaluate the role of this residue in catalysis. The mutant enzymes contained 1 mol of PLP per mol of subunit suggesting that Schiff base formation with lysine is not essential for PLP binding. The 3D structure of the mutant enzymes revealed that PLP was bound at the active site in an orientation different from that of the wild-type enzyme. In the presence of substrate, the PLP ring was in an orientation superimposable with that of the external aldimine complex of wild-type enzyme. However, the mutant enzymes were inactive, and the kinetic analysis of the different steps of catalysis revealed that there was a drastic reduction in the rate of formation of the quinonoid intermediate. Analysis of these results along with the crystal structures suggested that K-226 is responsible for flipping of PLP from one orientation to another which is crucial for H(4)PteGlu-dependent Calpha-Cbeta bond cleavage of l-Ser.
About this Structure
1YJS is a Protein complex structure of sequences from Geobacillus stearothermophilus with as ligand. Active as Glycine hydroxymethyltransferase, with EC number 2.1.2.1 Full crystallographic information is available from OCA.
Reference
Role of Lys-226 in the catalytic mechanism of Bacillus stearothermophilus serine hydroxymethyltransferase--crystal structure and kinetic studies., Bhavani S, Trivedi V, Jala VR, Subramanya HS, Kaul P, Prakash V, Appaji Rao N, Savithri HS, Biochemistry. 2005 May 10;44(18):6929-37. PMID:15865438
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