Proton linkage of complex formation between cytochrome c and cytochrome b5: electrostatic consequences of protein-protein interactions

Mr, Mauk; Pd, Barker; Ag, Mauk

doi:10.1021/bi00105a010

Cited by 38 publications

(37 citation statements)

References 58 publications

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“…These data suggest that electrostatic forces are involved in complex formation and that as solvent is excluded from the protein-protein interface hydration forces are replaced by salt linkages (Rogers, Pochapsky & Sligar, 1988;Mauk, Barker & Mauk, 1991;Kornblatt, Kornblatt, Hoa & Mauk, 1993). Computer models of such complexes (Salemme, 1976;Poulos & Mauk, 1983;Wendoloski, Mathew, Weber & Salemme, 1987), together with mutagenic studies, have identified negatively charged groups on b5 which link with positively charged residues surrounding the heme pocket of its partners.…”

Section: Discussionmentioning

confidence: 99%

Refinement and structural analysis of bovine cytochrome b5 at 1.5 Å Resolution

Durley¹,

Mathews²

1996

Acta Cryst D

146

198

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The structure of bovine liver cytochrome bs, a soluble 93-residue proteolytic fragment of a 16 kDa. membranebound hemoprotein, initially solved at 2.0A resolution, has been refined at 1.5/~ using data collected on a diffractometer. Refinement to 2.0/~ resolution used the Hendrickson-Konnert procedure PROLSQ and was then extended to 1.5 ~ resolution using the program PROFFT. Only residues 3-87 could be identified in the model and these residues together with 93 water molecules gave an agreement factor of R = 0.161 for data in the resolution range 1.5-5/~,. The structure was finally refined using the program X-PLOR, which enabled alternate conformers to be modelled for several surface side chains. Residues 1 and 2 at the amino terminus of the protein and residue 88 near the carboxyl terminus could be identified from these electron-density maps. However the remaining disordered carboxy-terminal residues could not successfully be included in the model. A total of 117 solvent molecules were included in the final refinement to give R =0.164 for the data between 1.5 and 10/~.

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Section: Discussionmentioning

confidence: 99%

Refinement and structural analysis of bovine cytochrome b5 at 1.5 Å Resolution

Durley¹,

Mathews²

1996

Acta Cryst D

146

198

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“…Binding of Mn 2ϩ ions to wild-type Mb and the variant was studied by monitoring the release of protons upon metal ion binding. Instrumentation (27), preparation of metal ion solutions (28), and data acquisition and analysis procedures (28,29) were reported previously. The dependence of peroxidase activity on pH was evaluated at 25°C between pH 5.0 and 8.0 with a mixed buffer system (5 mM Mes͞5 mM Mops͞5 mM TAPS͞95 mM NaCl) [ (32).…”

Section: Methodsmentioning

confidence: 99%

Introduction and characterization of a functionally linked metal ion binding site at the exposed heme edge of myoglobin

Hunter

Maurus²,

Mauk³

et al. 2003

Proc. Natl. Acad. Sci. U.S.A.

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A binding site for metal ions has been created on the surface of horse heart myoglobin (Mb) near the heme 6-propionate group by replacing K45 and K63 with glutamyl residues. One-dimensional 1 H NMR spectroscopy indicates that Mn 2؉ binds in the vicinity of the heme 6-propionate as anticipated, and potentiometric titrations establish that the affinity of the new site for Mn 2؉ is 1.28(4) ؋ 10 4 M ؊1 (pH 6.96, ionic strength I ‫؍‬ 17.2 M, 25°C). In addition, these substitutions lower the reduction potential of the protein and increase the pK a for the water molecule coordinated to the heme iron of metmyoglobin. The peroxidase [2,2 -azinobis(3-ethylbenzthiazoline-6-sulfonic acid), ABTS, as substrate] and the Mn 2؉ -peroxidase activity of the variant are both increased Ϸ3-fold. In contrast to wild-type Mb, both the affinity for azide and the midpoint potential of the variant are significantly influenced by the addition of Mn 2؉ . The structure of the variant has been determined by x-ray crystallography to define the coordination environment of bound Mn 2؉ and Cd 2؉ . Although slight differences are observed between the geometry of the binding of the two metal ions, both are hexacoordinate, and neither involves coordination by E63.

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“…The change in proton binding that is associated with the binding of Zn 2+ , Co 2+ , Ni 2+ , Mn 2+ , Cu 2+ and Fe 3+ to apoFurC* (5.69-6.44 µM) was studied at 25 • C by the method of Laskowski and Finkenstadt [37] with a Radiometer ABU93 Triburet operated under computer control as described previously [38]. Difficulties related to oxidation of Fe 2+ by dissolved dioxygen prevented consideration of the binding of this species to FurC*.…”

Section: Potentiometric Titrationsmentioning

confidence: 99%

Functional characterization of the dimerization domain of the ferric uptake regulator (Fur) of Pseudomonas aeruginosa

Bai

Rosell

Lige

et al. 2006

Biochemical Journal

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The functional properties of the recombinant C-terminal dimerization domain of the Pseudomonas aeruginosa Fur (ferric uptake regulator) protein expressed in and purified from Escherichia coli have been evaluated. Sedimentation velocity measurements demonstrate that this domain is dimeric, and the UV CD spectrum is consistent with a secondary structure similar to that observed for the corresponding region of the crystallographically characterized wild-type protein. The thermal stability of the domain as determined by CD spectroscopy decreases significantly as pH is increased and increases significantly as metal ions are added. Potentiometric titrations (pH 6.5) establish that the domain possesses a high-affinity and a low-affinity binding site for metal ions. The high-affinity (sensory) binding site demonstrates association constants (K(A)) of 10(+/-7)x10(6), 5.7(+/-3)x10(6), 2.0(+/-2)x10(6) and 2.0(+/-3)x10(4) M(-1) for Ni2+, Zn2+, Co2+ and Mn2+ respectively, while the low-affinity (structural) site exhibits association constants of 1.3(+/-2)x10(6), 3.2(+/-2)x10(4), 1.76(+/-1)x10(5) and 1.5(+/-2)x10(3) M(-1) respectively for the same metal ions (pH 6.5, 300 mM NaCl, 25 degrees C). The stability of metal ion binding to the sensory site follows the Irving-Williams order, while metal ion binding to the partial sensory site present in the domain does not. Fluorescence experiments indicate that the quenching resulting from binding of Co2+ is reversed by subsequent titration with Zn2+. We conclude that the domain is a reasonable model for many properties of the full-length protein and is amenable to some analyses that the limited solubility of the full-length protein prevents.

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Proton linkage of complex formation between cytochrome c and cytochrome b5: electrostatic consequences of protein-protein interactions

Cited by 38 publications

References 58 publications

Refinement and structural analysis of bovine cytochrome b5 at 1.5 Å Resolution

Refinement and structural analysis of bovine cytochrome b5 at 1.5 Å Resolution

Introduction and characterization of a functionally linked metal ion binding site at the exposed heme edge of myoglobin

Functional characterization of the dimerization domain of the ferric uptake regulator (Fur) of Pseudomonas aeruginosa

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