1d5h: Difference between revisions

Revision as of 13:13, 21 February 2008

Rnase s(f8a). mutant ribonucleasE S.

OverviewOverview

The hydrophobic effect is widely believed to be an important determinant of protein stability. However, it is difficult to obtain unambiguous experimental estimates of the contribution of the hydrophobic driving force to the overall free energy of folding. Thermodynamic and structural studies of large to small substitutions in proteins are the most direct method of measuring this contribution. We have substituted the buried residue Phe8 in RNase S with alanine, methionine, and norleucine. Binding thermodynamics and structures were characterized by titration calorimetry and crystallography, respectively. The crystal structures of the RNase S F8A, F8M, and F8Nle mutants indicate that the protein tolerates the changes without any main chain adjustments. The correlation of structural and thermodynamic parameters associated with large to small substitutions was analyzed for nine mutants of RNase S as well as 32 additional cavity-containing mutants of T4 lysozyme, human lysozyme, and barnase. Such substitutions were typically found to result in negligible changes in DeltaC(p)() and positive values of both DeltaDeltaH degrees and DeltaDeltaS of folding. Enthalpic effects were dominant, and the sign of DeltaDeltaS is the opposite of that expected from the hydrophobic effect. Values of DeltaDeltaG degrees and DeltaDeltaH degrees correlated better with changes in packing parameters such as residue depth or occluded surface than with the change in accessible surface area upon folding. These results suggest that the loss of packing interactions rather than the hydrophobic effect is a dominant contributor to the observed energetics for large to small substitutions. Hence, estimates of the magnitude of the hydrophobic driving force derived from earlier mutational studies are likely to be significantly in excess of the actual value.

About this StructureAbout this Structure

1D5H is a Protein complex structure of sequences from Bos taurus with as ligand. Full crystallographic information is available from OCA.

ReferenceReference

Thermodynamic and structural studies of cavity formation in proteins suggest that loss of packing interactions rather than the hydrophobic effect dominates the observed energetics., Ratnaparkhi GS, Varadarajan R, Biochemistry. 2000 Oct 10;39(40):12365-74. PMID:11015216

Page seeded by OCA on Thu Feb 21 12:13:12 2008

Proteopedia Page Contributors and Editors (what is this?)Proteopedia Page Contributors and Editors (what is this?)

OCA

@@ Line 1: / Line 1: @@
-[[Image:1d5h.gif|left|200px]]<br /><applet load="1d5h" size="450" color="white" frame="true" align="right" spinBox="true"
+[[Image:1d5h.gif|left|200px]]<br /><applet load="1d5h" size="350" color="white" frame="true" align="right" spinBox="true"
 caption="1d5h, resolution 2.25&Aring;" />
 '''Rnase s(f8a). mutant ribonucleasE S.'''<br />
 ==Overview==
-The hydrophobic effect is widely believed to be an important determinant, of protein stability. However, it is difficult to obtain unambiguous, experimental estimates of the contribution of the hydrophobic driving, force to the overall free energy of folding. Thermodynamic and structural, studies of large to small substitutions in proteins are the most direct, method of measuring this contribution. We have substituted the buried, residue Phe8 in RNase S with alanine, methionine, and norleucine. Binding, thermodynamics and structures were characterized by titration calorimetry, and crystallography, respectively. The crystal structures of the RNase S, F8A, F8M, and F8Nle mutants indicate that the protein tolerates the, changes without any main chain adjustments. The correlation of structural, and thermodynamic parameters associated with large to small substitutions, was analyzed for nine mutants of RNase S as well as 32 additional, cavity-containing mutants of T4 lysozyme, human lysozyme, and barnase., Such substitutions were typically found to result in negligible changes in, DeltaC(p)() and positive values of both DeltaDeltaH degrees and, DeltaDeltaS of folding. Enthalpic effects were dominant, and the sign of, DeltaDeltaS is the opposite of that expected from the hydrophobic effect., Values of DeltaDeltaG degrees and DeltaDeltaH degrees correlated better, with changes in packing parameters such as residue depth or occluded, surface than with the change in accessible surface area upon folding., These results suggest that the loss of packing interactions rather than, the hydrophobic effect is a dominant contributor to the observed, energetics for large to small substitutions. Hence, estimates of the, magnitude of the hydrophobic driving force derived from earlier mutational, studies are likely to be significantly in excess of the actual value.
+The hydrophobic effect is widely believed to be an important determinant of protein stability. However, it is difficult to obtain unambiguous experimental estimates of the contribution of the hydrophobic driving force to the overall free energy of folding. Thermodynamic and structural studies of large to small substitutions in proteins are the most direct method of measuring this contribution. We have substituted the buried residue Phe8 in RNase S with alanine, methionine, and norleucine. Binding thermodynamics and structures were characterized by titration calorimetry and crystallography, respectively. The crystal structures of the RNase S F8A, F8M, and F8Nle mutants indicate that the protein tolerates the changes without any main chain adjustments. The correlation of structural and thermodynamic parameters associated with large to small substitutions was analyzed for nine mutants of RNase S as well as 32 additional cavity-containing mutants of T4 lysozyme, human lysozyme, and barnase. Such substitutions were typically found to result in negligible changes in DeltaC(p)() and positive values of both DeltaDeltaH degrees and DeltaDeltaS of folding. Enthalpic effects were dominant, and the sign of DeltaDeltaS is the opposite of that expected from the hydrophobic effect. Values of DeltaDeltaG degrees and DeltaDeltaH degrees correlated better with changes in packing parameters such as residue depth or occluded surface than with the change in accessible surface area upon folding. These results suggest that the loss of packing interactions rather than the hydrophobic effect is a dominant contributor to the observed energetics for large to small substitutions. Hence, estimates of the magnitude of the hydrophobic driving force derived from earlier mutational studies are likely to be significantly in excess of the actual value.
 ==About this Structure==
-D5H is a [http://en.wikipedia.org/wiki/Protein_complex Protein complex] structure of sequences from [http://en.wikipedia.org/wiki/Bos_taurus Bos taurus] with SO4 as [http://en.wikipedia.org/wiki/ligand ligand]. Full crystallographic information is available from [http://ispc.weizmann.ac.il/oca-bin/ocashort?id=1D5H OCA].
+D5H is a [http://en.wikipedia.org/wiki/Protein_complex Protein complex] structure of sequences from [http://en.wikipedia.org/wiki/Bos_taurus Bos taurus] with <scene name='pdbligand=SO4:'>SO4</scene> as [http://en.wikipedia.org/wiki/ligand ligand]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1D5H OCA].
 ==Reference==
@@ Line 13: / Line 13: @@
 [[Category: Bos taurus]]
 [[Category: Protein complex]]
-[[Category: Ratnaparkhi, G.S]]
+[[Category: Ratnaparkhi, G S]]
 [[Category: Varadarajan, R.]]
 [[Category: SO4]]
 [[Category: rnase s mutant f8a cavity s protein s peptide]]
-''Page seeded by [http://ispc.weizmann.ac.il/oca OCA ] on Tue Nov 20 13:01:19 2007''
+''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Feb 21 12:13:12 2008''