1e52
From Proteopedia
(New page: 200px<br /><applet load="1e52" size="450" color="white" frame="true" align="right" spinBox="true" caption="1e52" /> '''SOLUTION STRUCTURE OF ESCHERICHIA COLI UVRB ...) |
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'''SOLUTION STRUCTURE OF ESCHERICHIA COLI UVRB C-TERMINAL DOMAIN'''<br /> | '''SOLUTION STRUCTURE OF ESCHERICHIA COLI UVRB C-TERMINAL DOMAIN'''<br /> | ||
==Overview== | ==Overview== | ||
| - | The solution structure, thermodynamic stability and hydrodynamic | + | The solution structure, thermodynamic stability and hydrodynamic properties of the 55-residue C-terminal domain of UvrB that interacts with UvrC during excision repair in E. coli have been determined using a combination of high resolution NMR, ultracentrifugation, 15N NMR relaxation, gel permeation, NMR diffusion, circular dichroism and differential scanning calorimetry. The subunit molecular weight is 7,438 kDa., compared with 14.5+/-1.0 kDa. determined by equilibrium sedimentation, indicating a dimeric structure. The structure determined from NMR showed a stable dimer of anti-parallel helical hairpins that associate in an unusual manner, with a small and hydrophobic interface. The Stokes radius of the protein decreases from a high plateau value (ca. 22 A) at protein concentrations greater than 4 microM to about 18 A at concentrations less than 0.1 microM. The concentration and temperature-dependence of the far UV circular dichroism show that the protein is thermally stable (Tm ca. 71.5 degrees C at 36 microM). The simplest model consistent with these data was a dimer dissociating into folded monomers that then unfolds co-operatively. The van't Hoff enthalpy and dissociation constant for both transition was derived by fitting, with deltaH1=23 kJ mol(-1). K1(298)=0.4 microM and deltaH2= 184 kJ mol(-1). This is in good agreement with direct calorimetric analysis of the thermal unfolding of the protein, which gave a calorimetric enthalpy change of 181 kJ mol(-1) and a van't Hoff enthalpy change of 354 kJ mol(-1), confirming the dimer to monomer unfolding. The thermodynamic data can be reconciled with the observed mode of dimerisation. 15N NMR relaxation measurements at 14.1 T and 11.75 T confirmed that the protein behaves as an asymmetric dimer at mM concentrations, with a flexible N-terminal linker for attachment to the remainder of the UvrB protein. The role of dimerisation of this domain in the excision repair mechanism is discussed. |
==About this Structure== | ==About this Structure== | ||
| - | 1E52 is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Escherichia_coli Escherichia coli]. Full crystallographic information is available from [http:// | + | 1E52 is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Escherichia_coli Escherichia coli]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1E52 OCA]. |
==Reference== | ==Reference== | ||
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[[Category: Single protein]] | [[Category: Single protein]] | ||
[[Category: Alexandrovich, A.]] | [[Category: Alexandrovich, A.]] | ||
| - | [[Category: Frenkiel, T | + | [[Category: Frenkiel, T A.]] |
[[Category: Goosen, N.]] | [[Category: Goosen, N.]] | ||
[[Category: Kelly, G.]] | [[Category: Kelly, G.]] | ||
| - | [[Category: Lane, A | + | [[Category: Lane, A N.]] |
| - | [[Category: Moolenaar, G | + | [[Category: Moolenaar, G F.]] |
| - | [[Category: Sanderson, M | + | [[Category: Sanderson, M R.]] |
[[Category: dna repair]] | [[Category: dna repair]] | ||
[[Category: uvrb]] | [[Category: uvrb]] | ||
[[Category: uvrc binding domain]] | [[Category: uvrc binding domain]] | ||
| - | ''Page seeded by [http:// | + | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Feb 21 12:23:53 2008'' |
Revision as of 10:23, 21 February 2008
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SOLUTION STRUCTURE OF ESCHERICHIA COLI UVRB C-TERMINAL DOMAIN
Overview
The solution structure, thermodynamic stability and hydrodynamic properties of the 55-residue C-terminal domain of UvrB that interacts with UvrC during excision repair in E. coli have been determined using a combination of high resolution NMR, ultracentrifugation, 15N NMR relaxation, gel permeation, NMR diffusion, circular dichroism and differential scanning calorimetry. The subunit molecular weight is 7,438 kDa., compared with 14.5+/-1.0 kDa. determined by equilibrium sedimentation, indicating a dimeric structure. The structure determined from NMR showed a stable dimer of anti-parallel helical hairpins that associate in an unusual manner, with a small and hydrophobic interface. The Stokes radius of the protein decreases from a high plateau value (ca. 22 A) at protein concentrations greater than 4 microM to about 18 A at concentrations less than 0.1 microM. The concentration and temperature-dependence of the far UV circular dichroism show that the protein is thermally stable (Tm ca. 71.5 degrees C at 36 microM). The simplest model consistent with these data was a dimer dissociating into folded monomers that then unfolds co-operatively. The van't Hoff enthalpy and dissociation constant for both transition was derived by fitting, with deltaH1=23 kJ mol(-1). K1(298)=0.4 microM and deltaH2= 184 kJ mol(-1). This is in good agreement with direct calorimetric analysis of the thermal unfolding of the protein, which gave a calorimetric enthalpy change of 181 kJ mol(-1) and a van't Hoff enthalpy change of 354 kJ mol(-1), confirming the dimer to monomer unfolding. The thermodynamic data can be reconciled with the observed mode of dimerisation. 15N NMR relaxation measurements at 14.1 T and 11.75 T confirmed that the protein behaves as an asymmetric dimer at mM concentrations, with a flexible N-terminal linker for attachment to the remainder of the UvrB protein. The role of dimerisation of this domain in the excision repair mechanism is discussed.
About this Structure
1E52 is a Single protein structure of sequence from Escherichia coli. Full crystallographic information is available from OCA.
Reference
Solution structure, hydrodynamics and thermodynamics of the UvrB C-terminal domain., Alexandrovich A, Czisch M, Frenkiel TA, Kelly GP, Goosen N, Moolenaar GF, Chowdhry BZ, Sanderson MR, Lane AN, J Biomol Struct Dyn. 2001 Oct;19(2):219-36. PMID:11697728
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