1mnh: Difference between revisions

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INTERACTIONS AMONG RESIDUES CD3, E7, E10 AND E11 IN MYOGLOBINS: ATTEMPTS TO SIMULATE THE O2 AND CO BINDING PROPERTIES OF APLYSIA MYOGLOBIN

OverviewOverview

Site-directed mutations have been introduced singly and in combination at residues lysine/arginine45 (CD3), histidine64 (E7), threonine67 (E10), and valine68 (E11) in pig and sperm whale myoglobins. The mutations probe the roles of these key distal pocket residues and represent attempts to mimic the heme environment of Aplysia limacina myoglobin which achieves moderately high O2 affinity in the absence of a distal histidine. In the mollusc myoglobin, arginine-E10 is believed to swing into the heme pocket and provide a hydrogen bond to the bound O2. The association and dissociation rate constants for oxygen and carbon monoxide binding to H64V, T67A, T67V, T67E, T67R, V68I, V68T, H64V-T67R, H64V-V68T, H64V-V68I, and H64V-T67R-V68I pig myoglobin mutants and T67R, H64V-T67R, and R45D-H64V-T67R mutants of sperm whale myoglobin have been measured using stopped-flow rapid mixing and flash photolysis techniques. Replacement of histidine-E7 with valine in either pig or sperm whale myoglobin drastically lowers O2 affinity while increasing CO affinity. Two second-site mutations, T67R and V68T, increase O2 affinity in the H64V mutant, even though when introduced singly these mutations have no effect or lower KO2, respectively. However, the oxygen affinities of the H64V-T67R mutants are 5-10-fold lower than that of A. limacina myoglobin. The crystal structure of the pig H64V-T67R double mutant reveals that the valine-E7 side chain is approximately 1 A closer to the heme plane than in the mollusc protein which may restrict access of the arginine-E10 side chain into the heme pocket. The O2 affinity of the H64V-T67R double mutant is not altered by the R45D replacement but is reduced 10-fold by the V68I mutation. The interactive effects of the T67R, V68I, and V68T mutations with the H64V substitution are discussed in terms of O2, CO, and N3-binding and the crystal structures of the H64V-T67R, H64V-V68I, and H64V-V68T double-mutant proteins. In many instances, the effects of second-site mutations in the valine64 background are the opposite of those observed for the corresponding single mutations in the wild type background. These results can be understood in terms of the changes in the rate-determining steps for ligand association and dissociation and the loss of distal pocket water molecules which follow replacement of histidine64 by valine.

About this StructureAbout this Structure

1MNH is a Single protein structure of sequence from Sus scrofa with as ligand. Full crystallographic information is available from OCA.

ReferenceReference

Interactions among residues CD3, E7, E10, and E11 in myoglobins: attempts to simulate the ligand-binding properties of Aplysia myoglobin., Smerdon SJ, Krzywda S, Brzozowski AM, Davies GJ, Wilkinson AJ, Brancaccio A, Cutruzzola F, Allocatelli CT, Brunori M, Li T, et al., Biochemistry. 1995 Jul 11;34(27):8715-25. PMID:7612611

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 '''INTERACTIONS AMONG RESIDUES CD3, E7, E10 AND E11 IN MYOGLOBINS: ATTEMPTS TO SIMULATE THE O2 AND CO BINDING PROPERTIES OF APLYSIA MYOGLOBIN'''<br />
 ==Overview==
-Site-directed mutations have been introduced singly and in combination at, residues lysine/arginine45 (CD3), histidine64 (E7), threonine67 (E10), and, valine68 (E11) in pig and sperm whale myoglobins. The mutations probe the, roles of these key distal pocket residues and represent attempts to mimic, the heme environment of Aplysia limacina myoglobin which achieves, moderately high O2 affinity in the absence of a distal histidine. In the, mollusc myoglobin, arginine-E10 is believed to swing into the heme pocket, and provide a hydrogen bond to the bound O2. The association and, dissociation rate constants for oxygen and carbon monoxide binding to, H64V, T67A, T67V, T67E, T67R, V68I, V68T, H64V-T67R, H64V-V68T, H64V-V68I, and H64V-T67R-V68I pig myoglobin mutants and T67R, H64V-T67R, and, R45D-H64V-T67R mutants of sperm whale myoglobin have been measured using, stopped-flow rapid mixing and flash photolysis techniques. Replacement of, histidine-E7 with valine in either pig or sperm whale myoglobin, drastically lowers O2 affinity while increasing CO affinity. Two, second-site mutations, T67R and V68T, increase O2 affinity in the H64V, mutant, even though when introduced singly these mutations have no effect, or lower KO2, respectively. However, the oxygen affinities of the, H64V-T67R mutants are 5-10-fold lower than that of A. limacina myoglobin., The crystal structure of the pig H64V-T67R double mutant reveals that the, valine-E7 side chain is approximately 1 A closer to the heme plane than in, the mollusc protein which may restrict access of the arginine-E10 side, chain into the heme pocket. The O2 affinity of the H64V-T67R double mutant, is not altered by the R45D replacement but is reduced 10-fold by the V68I, mutation. The interactive effects of the T67R, V68I, and V68T mutations, with the H64V substitution are discussed in terms of O2, CO, and, N3-binding and the crystal structures of the H64V-T67R, H64V-V68I, and, H64V-V68T double-mutant proteins. In many instances, the effects of, second-site mutations in the valine64 background are the opposite of those, observed for the corresponding single mutations in the wild type, background. These results can be understood in terms of the changes in the, rate-determining steps for ligand association and dissociation and the, loss of distal pocket water molecules which follow replacement of, histidine64 by valine.
+Site-directed mutations have been introduced singly and in combination at residues lysine/arginine45 (CD3), histidine64 (E7), threonine67 (E10), and valine68 (E11) in pig and sperm whale myoglobins. The mutations probe the roles of these key distal pocket residues and represent attempts to mimic the heme environment of Aplysia limacina myoglobin which achieves moderately high O2 affinity in the absence of a distal histidine. In the mollusc myoglobin, arginine-E10 is believed to swing into the heme pocket and provide a hydrogen bond to the bound O2. The association and dissociation rate constants for oxygen and carbon monoxide binding to H64V, T67A, T67V, T67E, T67R, V68I, V68T, H64V-T67R, H64V-V68T, H64V-V68I, and H64V-T67R-V68I pig myoglobin mutants and T67R, H64V-T67R, and R45D-H64V-T67R mutants of sperm whale myoglobin have been measured using stopped-flow rapid mixing and flash photolysis techniques. Replacement of histidine-E7 with valine in either pig or sperm whale myoglobin drastically lowers O2 affinity while increasing CO affinity. Two second-site mutations, T67R and V68T, increase O2 affinity in the H64V mutant, even though when introduced singly these mutations have no effect or lower KO2, respectively. However, the oxygen affinities of the H64V-T67R mutants are 5-10-fold lower than that of A. limacina myoglobin. The crystal structure of the pig H64V-T67R double mutant reveals that the valine-E7 side chain is approximately 1 A closer to the heme plane than in the mollusc protein which may restrict access of the arginine-E10 side chain into the heme pocket. The O2 affinity of the H64V-T67R double mutant is not altered by the R45D replacement but is reduced 10-fold by the V68I mutation. The interactive effects of the T67R, V68I, and V68T mutations with the H64V substitution are discussed in terms of O2, CO, and N3-binding and the crystal structures of the H64V-T67R, H64V-V68I, and H64V-V68T double-mutant proteins. In many instances, the effects of second-site mutations in the valine64 background are the opposite of those observed for the corresponding single mutations in the wild type background. These results can be understood in terms of the changes in the rate-determining steps for ligand association and dissociation and the loss of distal pocket water molecules which follow replacement of histidine64 by valine.
 ==About this Structure==
-MNH is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Sus_scrofa Sus scrofa] with HEM as [http://en.wikipedia.org/wiki/ligand ligand]. Full crystallographic information is available from [http://ispc.weizmann.ac.il/oca-bin/ocashort?id=1MNH OCA].
+MNH is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Sus_scrofa Sus scrofa] with <scene name='pdbligand=HEM:'>HEM</scene> as [http://en.wikipedia.org/wiki/ligand ligand]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1MNH OCA].
 ==Reference==
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 [[Category: Single protein]]
 [[Category: Sus scrofa]]
-[[Category: Davies, G.J.]]
+[[Category: Davies, G J.]]
-[[Category: Wilkinson, A.J.]]
+[[Category: Wilkinson, A J.]]
 [[Category: HEM]]
 [[Category: oxygen storage]]
-''Page seeded by [http://ispc.weizmann.ac.il/oca OCA ] on Tue Nov 20 21:33:16 2007''
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