Structural perturbations of azurin deposited on solid matrices as revealed by trp phosphorescence.
The phosphorescence emission of Cd-azurin from Pseudomonas aeruginosa was used as a probe of possible perturbations in the dynamical structure of the protein core that may be induced by protein-sorbent and protein-protein interactions occurring when the macromolecule is deposited into amorphous, thin solid films. Relative to the protein in aqueous solution, the spectrum is unrelaxed and the phosphorescence decay becomes highly heterogeneous, the average lifetime increasing sharply with film thickness and upon its dehydration. According to the lifetime parameter, adsorption of the protein to the substrate is found to produce a multiplicity of partially unfolded structures, an influence that propagates for several protein layers from the surface. Among the substrates used for film deposition, hydrophilic silica, dextran, DEAE-dextran, dextran sulfate, and hydrophobic octodecylamine, the perturbation is smallest with dextran sulfate and largest with octodecylamine. The destabilizing effect of protein-protein interactions, as monitored on 50-layer-thick films, is most evident at a relative humidity of 75%. Stabilizing agents were incorporated to attenuate the deleterious effects of protein aggregation. Among them, the most effective in preserving a more native-like structure are the disaccharides sucrose and trehalose in dry films and the polymer dextran in wet films. Interestingly, the polymer was found to achieve maximum efficacy at sensibly lower additive/protein ratios than the sugars.
Gabellieri
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Consiglio Nazionale delle Ricerche, Istituto di Biofisica, Pisa, Italy.
Strambini
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GB
D000327:Adsorption; D001400:Azurin / Q000737:chemistry*; D002104:Cadmium / Q000737:chemistry; D003637:DEAE-Dextran / Q000737:chemistry; D016264:Dextran Sulfate / Q000737:chemistry; D003911:Dextrans / Q000737:chemistry; D049449:Luminescence; D011485:Protein Binding; D017510:Protein Folding; D011550:Pseudomonas aeruginosa / Q000737:chemistry; D012825:Silicon / Q000737:chemistry; D013050:Spectrometry, Fluorescence; D014364:Tryptophan / Q000737:chemistry*; D014867:Water / Q000378:metabolism
D003911:Dextrans; D002104:Cadmium; D014867:Water; D001400:Azurin; D014364:Tryptophan; D003637:DEAE-Dextran; D016264:Dextran Sulfate; D012825:Silicon