Gated Electron Transfers and Electron Pathways in Azurin: A NMR Dynamic Study at Multiple Fields and Temperatures

Zhuravleva, Anastasia; Korzhnev, Dmitry M.; Kupče, Ēriks; Arseniev, Alexander S.; Billeter, Martin; Orekhov, Vladislav Yu.

doi:10.1016/j.jmb.2004.08.001

Cited by 31 publications

(36 citation statements)

References 77 publications

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“…In order to extend these results to other cupredoxins, we have studied the solution dynamics of P. aeruginosa apo-azurin. The blue copper protein azurin has been extensively studied as a paradigmatic protein for electron transfer (42)(43)(44), and Cu(I)-azurin has been the subject of several NMR studies (55)(56)(57)(58). Instead, apo-azurin has not been characterized by NMR.…”

Section: Resultsmentioning

confidence: 99%

Flexibility of the metal-binding region in apo-cupredoxins

Zaballa

Abriata

Donaire

et al. 2012

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

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Protein-mediated electron transfer is an essential event in many biochemical processes. Efficient electron transfer requires the reorganization energy of the redox event to be minimized, which is ensured by the presence of rigid donor and acceptor sites. Electron transfer copper sites are present in the ubiquitous cupredoxin fold, able to bind one or two copper ions. The low reorganization energy in these metal centers has been accounted for by assuming that the protein scaffold creates an entatic/rack-induced state, which gives rise to a rigid environment by means of a preformed metal chelating site. However, this notion is incompatible with the need for an exposed metal-binding site and protein-protein interactions enabling metallochaperone-mediated assembly of the copper site. Here we report an NMR study that reveals a high degree of structural heterogeneity in the metal-binding region of the nonmetallated Cu A -binding cupredoxin domain, arising from microsecond to second dynamics that are quenched upon metal binding. We also report similar dynamic features in apo-azurin, a paradigmatic blue copper protein, suggesting a general behavior. These findings reveal that the entatic/rack-induced state, governing the features of the metal center in the copper-loaded protein, does not require a preformed metal-binding site. Instead, metal binding is a major contributor to the rigidity of electron transfer copper centers. These results reconcile the seemingly contradictory requirements of a rigid, occluded center for electron transfer, and an accessible, dynamic site required for in vivo copper uptake.metalloproteins | nuclear magnetic resonance | protein dynamics

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Section: Resultsmentioning

confidence: 99%

Flexibility of the metal-binding region in apo-cupredoxins

Zaballa

Abriata

Donaire

et al. 2012

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

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“…47 The spectra were processed and quantified using NMRPipe, 48 MUNIN, 49 and DASHA 50 software packages as described. 51 15 N relaxation measurements 15 N longitudinal (R 1 ) and transverse (R 2 ) relaxation rates were measured for free barnase and the barnasebarstar complex at 500 MHz (spectrometer 1 H Larmor frequency) using pulse sequences described previously. 52 Additionally, R 1 and R 2 experiments (600 MHz) were performed for a dilute barnase-barstar complex (0.17 mM concentration).…”

Section: Nmr Relaxation Experimentsmentioning

confidence: 99%

Propagation of Dynamic Changes in Barnase Upon Binding of Barstar: An NMR and Computational Study

Zhuravleva

Korzhnev

Nolde

et al. 2007

Journal of Molecular Biology

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“…[1][2][3] For instance, protein dynamics has been recently recognized as a key factor in controlling or limiting interand intraprotein ET reactions. [4][5][6][7][8][9][10][11][12][13][14][15] However, in most cases the complexity of the systems impairs direct observations of conformational gating, configurational fluctuations, or rearrangement of protein complexes under reactive conditions. In this context, the combination of suitable spectroelectrochemical techniques with simplified model systems, such as proteins immobilized on biomimetic electrodes, can greatly contribute to the elucidation of the biophysical fundamentals in better detail.…”

Section: Introductionmentioning

confidence: 99%

Thermal Fluctuations Determine the Electron‐Transfer Rates of Cytochrome c in Electrostatic and Covalent Complexes

Martí

Martín

et al. 2010

ChemPhysChem

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The heterogeneous electron-transfer (ET) reaction of cytochrome c (Cyt-c) electrostatically or covalently immobilized on electrodes coated with self-assembled monolayers (SAMs) of omega-functionalized alkanethiols is analyzed by surface-enhanced resonance Raman (SERR) spectroscopy and molecular dynamics (MD) simulations. Electrostatically bound Cyt-c on pure carboxyl-terminated and mixed carboxyl/hydroxyl-terminated SAMs reveals the same distance dependence of the rate constants, that is, electron tunneling at long distances and a regime controlled by the protein orientational distribution and dynamics that leads to a nearly distance-independent rate constant at short distances. Qualitatively, the same behavior is found for covalently bound Cyt-c, although the apparent ET rates in the plateau region are lower since protein mobility is restricted due to formation of amide bonds between the protein and the SAM. The experimental findings are consistent with the results of MD simulations indicating that thermal fluctuations of the protein and interfacial solvent molecules can effectively modulate the electron tunneling probability.

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Gated Electron Transfers and Electron Pathways in Azurin: A NMR Dynamic Study at Multiple Fields and Temperatures

Cited by 31 publications

References 77 publications

Flexibility of the metal-binding region in apo-cupredoxins

Flexibility of the metal-binding region in apo-cupredoxins

Propagation of Dynamic Changes in Barnase Upon Binding of Barstar: An NMR and Computational Study

Thermal Fluctuations Determine the Electron‐Transfer Rates of Cytochrome c in Electrostatic and Covalent Complexes

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