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43973-GB4
Using a Photolabile Precursor to Study Radical-Mediated Protein Damage

K. Nolan Carter, University of Central Arkansas

            Oxidative damage of proteins is believed to involve free radical intermediates.  This damage process can be initiated by reactive oxygen species such as hydroxyl radical.  Protein radicals thus produced can undergo subsequent reactions which are often complex and may result in transfer of damage to other biological molecules such as nucleic acids.  A problem associated with the direct study of this damage process is that hydroxyl radical is highly reactive and can produce multiple radicals from a given amino acid within a polypeptide chain.  Since this is possible at every amino acid within a polypeptide, many protein radicals can result from attack of hydroxyl radical.  This hinders direct study of specific protein radical intermediates involved in this process.  The objective of this research was to develop photochemically active amino acids which generate the same radicals produced by hydroxyl radical, but in a selective fashion.  It was envisioned that these photochemically active amino acids would contain a photolabile group which would be cleaved upon photolysis to generate amino acid radicals.  Radicals such as 1, derived from the reaction of hydroxyl radical with valine can be selectively generated under mild conditions via this approach.  We have successfully produced a photolabile precursor for a primary valine radical, the first such example of a compound of this type.

            Since alkyl aryl selenides are known to undergo facile C-Se bond homolysis upon UV irradiation, the phenylselenyl group was chosen as a photolabile group.  Phthaloyl protected diastereomeric valine radical precursors 4 and 5 were prepared from the corresponding diastereomeric bromides (2,3) by treatment with benzeneselenol in the presence of cesium carbonate.  This method is superior to more conventional methods for introduction of the phenylselenyl group such as diphenyl diselenide/sodium borohydride.  Removal of the phthaloyl protecting group with methanolic hydrazine afforded radical precursors 6 and 7 as a 2:1 mixture of diastereomers.  This compound was intended to generate a monomeric valine radical that would serve as a model for the analogous valine radical produced within proteins by hydroxyl radical. 

            Upon photolysis at 350 nm, this compound is converted to products consistent with the intermediacy of radicals 8 and 9.  Detailed characterization of photolysates by BSTFA derivatization followed by GC/MS analysis is in progress.

            Studies toward the synthesis of tertiary radical precursor 10 have also been conducted.  This compound is intended to produce tertiary valine radical 11.

            Due to the scarcity of available methods for the synthesis of tertiary alkyl aryl selenides, synthesis of tertiary radical precursor 10 has yet to be completed.  Having attempted unsuccessfully to prepare this compound from substitution of the corresponding alkyl halide, we are currently attempting to complete the synthesis via 1,4-addition of benzeneselenolate to alkene 12 to produce protected radical precursor 13.

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