Photodynamically generated bovine serum albumin radicals: evidence for damage transfer and oxidation at cysteine and tryptophan residues

TY - JOUR

T1 - Photodynamically generated bovine serum albumin radicals

T2 - evidence for damage transfer and oxidation at cysteine and tryptophan residues

AU - Silvester, J A

AU - Timmins, G S

AU - Davies, Michael Jonathan

PY - 1998/3/15

Y1 - 1998/3/15

N2 - Porphyrin-sensitized photoxidation of bovine serum albumin (BSA) results in oxidation of the protein at (at least) two different, specific sites: the Cys-34 residue giving rise to a thiyl radical (RS.); and one or both of the tryptophan residues (Trp-134 and Trp-214) resulting in the formation of tertiary carbon-centred radicals and disruption of the tryptophan ring system. In the case of porphyrins such as hematoporphyrin, which bind at specific sites on BSA, these species appear to arise via long-range transfer of damage within the protein structure, as the binding site is some distance from the ultimate site of radical formation. This transfer of damage is shown to depend on a number of factors including the conformation of the protein, the presence of blocking groups and pH. Alteration of the protein conformation results in radical formation at additional (or alternative) sites, as does blocking of the preferred loci of radical formation. The formation of these thiyl and tryptophan-derived radicals does not lead to significant aggregation or fragmentation of the protein, though it does result in a dramatic enhancement in the susceptibility of the oxidised protein to proteolytic degradation by a range of proteases. The generation of protein-derived radicals also results in an enhancement of photobleaching of the porphyrin, suggesting that protein radical generation is linked to porphyrin photooxidation.

AB - Porphyrin-sensitized photoxidation of bovine serum albumin (BSA) results in oxidation of the protein at (at least) two different, specific sites: the Cys-34 residue giving rise to a thiyl radical (RS.); and one or both of the tryptophan residues (Trp-134 and Trp-214) resulting in the formation of tertiary carbon-centred radicals and disruption of the tryptophan ring system. In the case of porphyrins such as hematoporphyrin, which bind at specific sites on BSA, these species appear to arise via long-range transfer of damage within the protein structure, as the binding site is some distance from the ultimate site of radical formation. This transfer of damage is shown to depend on a number of factors including the conformation of the protein, the presence of blocking groups and pH. Alteration of the protein conformation results in radical formation at additional (or alternative) sites, as does blocking of the preferred loci of radical formation. The formation of these thiyl and tryptophan-derived radicals does not lead to significant aggregation or fragmentation of the protein, though it does result in a dramatic enhancement in the susceptibility of the oxidised protein to proteolytic degradation by a range of proteases. The generation of protein-derived radicals also results in an enhancement of photobleaching of the porphyrin, suggesting that protein radical generation is linked to porphyrin photooxidation.

KW - Bicyclo Compounds, Heterocyclic

KW - Cysteine

KW - Free Radicals

KW - Hematoporphyrins

KW - Hydrogen-Ion Concentration

KW - Oxidation-Reduction

KW - Phosphites

KW - Photic Stimulation

KW - Photosensitizing Agents

KW - Serum Albumin, Bovine

KW - Spectrometry, Fluorescence

KW - Spin Trapping

KW - Tryptophan

M3 - Journal article

C2 - 9586806

SN - 0891-5849

VL - 24

SP - 754

EP - 766

JO - Free Radical Biology & Medicine

JF - Free Radical Biology & Medicine

IS - 5

ER -