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Electrochemical and structural properties of a protein system designed to generate tyrosine Pourbaix diagrams.

ABSTRACT: This report describes a model protein specifically tailored to electrochemically study the reduction potential of protein tyrosine radicals as a function of pH. The model system is based on the 67-residue ?(3)Y three-helix bundle. ?(3)Y contains a single buried tyrosine at position 32 and displays structural properties inherent to a protein. The present report presents differential pulse voltammograms obtained from ?(3)Y at both acidic (pH 5.6) and alkaline (pH 8.3) conditions. The observed Faradaic response is uniquely associated with Y32, as shown by site-directed mutagenesis. This is the first time voltammetry is successfully applied to detect a redox-active tyrosine residing in a structured protein environment. Tyrosine is a proton-coupled electron-transfer cofactor making voltammetry-based pH titrations a central experimental approach. A second set of experiments was performed to demonstrate that pH-dependent studies can be conducted on the redox-active tyrosine without introducing large-scale structural changes in the protein scaffold. ?(3)Y was re-engineered with the specific aim to place the imidazole group of a histidine close to the Y32 phenol ring. ?(3)Y-K29H and ?(3)Y-K36H each contain a histidine residue whose protonation perturbs the fluorescence of Y32. We show that these variants are stable and well-folded proteins whose helical content, tertiary structure, solution aggregation state, and solvent-sequestered position of Y32 remain pH insensitive across a range of at least 3-4 pH units. These results confirm that the local environment of Y32 can be altered and the resulting radical site studied by voltammetry over a broad pH range without interference from long-range structural effects.

SUBMITTER: Martinez-Rivera MC

PROVIDER: S-EPMC4076202 | biostudies-literature | 2011 Nov

REPOSITORIES: biostudies-literature

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Electrochemical and structural properties of a protein system designed to generate tyrosine Pourbaix diagrams.

Martínez-Rivera Melissa C MC Berry Bruce W BW Valentine Kathleen G KG Westerlund Kristina K Hay Sam S Tommos Cecilia C

Journal of the American Chemical Society 20111019 44

This report describes a model protein specifically tailored to electrochemically study the reduction potential of protein tyrosine radicals as a function of pH. The model system is based on the 67-residue α(3)Y three-helix bundle. α(3)Y contains a single buried tyrosine at position 32 and displays structural properties inherent to a protein. The present report presents differential pulse voltammograms obtained from α(3)Y at both acidic (pH 5.6) and alkaline (pH 8.3) conditions. The observed Fara ...[more]

PMID: 22011192

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Project description:Rare-earth labeling in biological apatite could provide critical information for the pathologic transition (osteoclastic) and physiologic regeneration (osteogenesis) of bone and teeth because of their characteristic site-sensitive fluorescence in different coordinative conditions of various tissues in many biological processes. However, the rare-earth labeling method for biological apatites, i.e., carbonated-hydroxyapatite, has been rarely found in the literature. In this paper, we report a Pourbaix-diagram guided mineralizing strategy to controllable carbonation and doping of rare-earth ions in the hydroxyapatite (HA) lattice. The carbonation process of hydroxyapatite was achieved by controllable mineralization in hydrothermal condition with K2CO3 as the carbonate source, which results into the pure B-type carbonated hydroxyapatite (CHA) with tunable carbonate substitution degree. All of the as-synthesized materials crystalized into P63/m (No. 176) space group with the lattice parameter of a decreases and c increases with the increasing of carbonate content in the reactants. Structural refinement results revealed that the substitution of planar CO32- is superimposed on one of the faces of PO43- tetrahedral sub-units with a rotation angle of 30° in reference to c-axis. All of the hydrothermally synthesized CHA nanocrystals show hexagonal rod-like morphology with the length of 70-110 nm and diameter of 21-35 nm, and the decreasing length/diameter ratio from 3.61 to 2.96 from low to high carbonated level of the samples. Five rare-earth cations, of Pr3+, Sm3+, Eu3+, Tb3+, and Ho3+, were used as possible probe ions that can be doped into either HA or CHA lattice. The site-preference of Tb3+ doping is the same in the crystallographic site of HA and CHA according to characteristic emission peaks of 5D4-7Fj (j = 3-6) transitions in their photoluminescent spectroscopy. Our work provides a controllable carbonation method for rare-earth labeling hydroxyapatite nanomaterials with potential biologically active implant powders for bone repair and tissue regeneration.

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Electrochemical and structural properties of a protein system designed to generate tyrosine Pourbaix diagrams.

Publications

Electrochemical and structural properties of a protein system designed to generate tyrosine Pourbaix diagrams.

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