2p3r
From Proteopedia
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==Overview== | ==Overview== | ||
| - | The crystal structure of an Escherichia coli glycerol kinase mutant Gly230 | + | The crystal structure of an Escherichia coli glycerol kinase mutant Gly230 --> Asp (GKG230D) was determined to 2.0 A resolution using a microfluidics based crystallization platform. The crystallization strategy involved a suite of microfluidic devices that characterized the solubility trends of GKG230D, performed nanoliter volume free interface diffusion crystallization experiments, and produced diffraction-quality crystals for in situ data collection. GKG230D displays increased enzymatic activity and decreased allosteric regulation by the glycolytic pathway intermediate fructose 1,6-bisphosphate (FBP) compared to wild-type GK (GKWT). Structural analysis revealed that the decreased allosteric regulation is a result of the altered FBP binding loop conformations in GKG230D that interfere with the wild-type FBP binding site. The altered FBP binding loop conformations in GKG230D are supported through a series of intramolecular loop interactions. The appearance of Asp230 in the FBP binding loops also repositions the wild-type FBP binding residues away from the FBP binding site. Light scattering analysis confirmed GKG230D is a dimer and is resistant to tetramer formation in the presence of FBP, whereas GKWT dimers are converted into putatively inactive tetramers in the presence of FBP. GKG230D also provides the first structural evidence for multiple GK monomer conformations in the presence of glycerol and in the absence of a nucleotide substrate and verifies that glycerol binding is not responsible for locking GK into the closed conformation necessary for GK activity. |
==About this Structure== | ==About this Structure== | ||
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==Reference== | ==Reference== | ||
| - | Crystal | + | Crystal structure of a hyperactive Escherichia coli glycerol kinase mutant Gly230 --> Asp obtained using microfluidic crystallization devices., Anderson MJ, DeLabarre B, Raghunathan A, Palsson BO, Brunger AT, Quake SR, Biochemistry. 2007 May 15;46(19):5722-31. Epub 2007 Apr 19. PMID:[http://ispc.weizmann.ac.il//pmbin/getpm?pmid=17441732 17441732] |
[[Category: Escherichia coli]] | [[Category: Escherichia coli]] | ||
[[Category: Glycerol kinase]] | [[Category: Glycerol kinase]] | ||
[[Category: Single protein]] | [[Category: Single protein]] | ||
| - | [[Category: Anderson, M | + | [[Category: Anderson, M J.]] |
| - | [[Category: Brunger, A | + | [[Category: Brunger, A T.]] |
[[Category: DeLaBarre, B.]] | [[Category: DeLaBarre, B.]] | ||
| - | [[Category: Quake, S | + | [[Category: Quake, S R.]] |
[[Category: GOL]] | [[Category: GOL]] | ||
[[Category: allosteric regulation]] | [[Category: allosteric regulation]] | ||
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[[Category: microfluidics]] | [[Category: microfluidics]] | ||
| - | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on | + | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Feb 21 18:25:35 2008'' |
Revision as of 16:25, 21 February 2008
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Crystal structure of a hyperactive Escherichia coli glycerol kinase mutant Gly230->Asp obtained using microfluidic crystallization devices
Overview
The crystal structure of an Escherichia coli glycerol kinase mutant Gly230 --> Asp (GKG230D) was determined to 2.0 A resolution using a microfluidics based crystallization platform. The crystallization strategy involved a suite of microfluidic devices that characterized the solubility trends of GKG230D, performed nanoliter volume free interface diffusion crystallization experiments, and produced diffraction-quality crystals for in situ data collection. GKG230D displays increased enzymatic activity and decreased allosteric regulation by the glycolytic pathway intermediate fructose 1,6-bisphosphate (FBP) compared to wild-type GK (GKWT). Structural analysis revealed that the decreased allosteric regulation is a result of the altered FBP binding loop conformations in GKG230D that interfere with the wild-type FBP binding site. The altered FBP binding loop conformations in GKG230D are supported through a series of intramolecular loop interactions. The appearance of Asp230 in the FBP binding loops also repositions the wild-type FBP binding residues away from the FBP binding site. Light scattering analysis confirmed GKG230D is a dimer and is resistant to tetramer formation in the presence of FBP, whereas GKWT dimers are converted into putatively inactive tetramers in the presence of FBP. GKG230D also provides the first structural evidence for multiple GK monomer conformations in the presence of glycerol and in the absence of a nucleotide substrate and verifies that glycerol binding is not responsible for locking GK into the closed conformation necessary for GK activity.
About this Structure
2P3R is a Single protein structure of sequence from Escherichia coli with as ligand. Active as Glycerol kinase, with EC number 2.7.1.30 Full crystallographic information is available from OCA.
Reference
Crystal structure of a hyperactive Escherichia coli glycerol kinase mutant Gly230 --> Asp obtained using microfluidic crystallization devices., Anderson MJ, DeLabarre B, Raghunathan A, Palsson BO, Brunger AT, Quake SR, Biochemistry. 2007 May 15;46(19):5722-31. Epub 2007 Apr 19. PMID:17441732
Page seeded by OCA on Thu Feb 21 18:25:35 2008
