Light-induced conversion of Trp to Gly and Gly hydroperoxide in IgG1.

The exposure of IgG1 in aqueous solution to light with λ = 254 nm or λ > 295 nm yields products consistent with Trp radical cation formation followed by (α)C-(ß)C cleavage of the Trp side chain. The resulting glycyl radicals either are reduced to Gly or add oxygen prior to reduction to Gly hydroperoxide. Photoirradiation at λ = 254 nm targets Trp at positions 191 (light chain), 309 and 377 (heavy chain) while photoirradiation at λ > 295 nm targets Trp at position 309 (heavy chain). Mechanistically, the formation of Trp radical cations likely proceeds via photoinduced electron or hydrogen transfer to disulfide bonds, yielding thiyl radicals and thiols, where thiols may serve as reductants for the intermediary glycyl or glycylperoxyl radicals.

Photolysis of recombinant human insulin in the solid state: formation of a dithiohemiacetal product at the C-terminal disulfide bond.

PURPOSE: Exposure of protein pharmaceuticals to light can result in chemical and physical modifications, potentially leading to loss of potency, aggregation, and/or immunogenicity. To correlate these potential consequences with molecular changes, the nature of photoproducts and their mechanisms of formation must be characterized. The present study focuses on the photochemical degradation of insulin in the solid state. METHODS: Solid insulin was characterized by solid-state NMR, polarized optical microscopy and scanning electron microscopy; various insulin preparations were exposed to UV light prior to product analysis by mass spectrometry. RESULTS: UV-exposure of solid human insulin results in photodissociation of the C-terminal intrachain disulfide bond, leading to formation of a CysS(•) thiyl radical pair which ultimately disproportionates into thiol and thioaldehyde species. The high reactivity of the thioaldehyde and proximity to the thiol allow the formation of a dithiohemiacetal structure. Dithiohemiacetal is formed during the UV-exposure of both crystalline and amorphous insulin. CONCLUSIONS: Dithiohemiacetals represent novel structures generated through the photochemical modification of disulfide bonds. This is the first time that such structure is identified during the photolysis of a protein in the solid state.

Selenium-modified TiO2 and its impact on photocatalysis.

This work describes the preparation of a selenium-modified TiO(2) photocatalyst and a preliminary evaluation of its photocatalytic activity. Se-TiO(2) displayed greater visible absorption than undoped TiO(2) and was still capable of degrading quinoline at a slightly faster rate than undoped TiO(2) under UV light. Se-TiO(2) was also able to degrade organic molecules under purely visible light by a single electron transfer pathway. Irradiation with >435 nm light showed no evidence of efficient production of HO•-like species. Se-TiO(2) was also examined under hypoxic conditions, where the Se atoms were capable of trapping photogenerated electrons as evidenced by XPS.

Toward the physiological basis for increased Agrotis ipsilon multiple nucleopolyhedrovirus infection following feeding of Agrotis ipsilon larvae on transgenic corn expressing Cry1Fa2.

Larvae of the black cutworm, Agrotis ipsilon Hufnagel, were more susceptible to infection by A. ipsilon multiple nucleopolyhedrovirus (AgipMNPV: Baculoviridae) after feeding on Herculex I, a transgenic corn hybrid expressing the Bacillus thuringiensis (Bt)-derived toxin Cry1Fa2 compared to larvae fed on isoline corn. We investigated the physiological basis for increased susceptibility to virus infection following exposure to Herculex I by analyzing the midgut pH, gut protease activity and peritrophic matrix structure which are important factors for both Bt toxin action and baculovirus infection. No significant treatment differences were found in the pH of anterior midgut, central midgut or posterior midgut in larvae fed Herculex I or isoline diets. Analysis of soluble and membrane-associated gut proteinase activities from larvae fed Herculex I or isoline diets indicated that membrane-associated aminopeptidase activity and soluble chymotrypsin-like proteinase activity were significantly lower in Herculex I -fed larvae compared to isoline-fed larvae. The number and relative molecular masses of soluble chymotrypsin-like proteinases did not differ. Baculoviruses were not susceptible to in vitro degradation by bovine chymotrypsin, suggesting that chymotrypsin degradation of baculovirus occlusion-derived virus did not result in reduced infection of larvae fed on isoline diet. Scanning electron micrographs of the peritrophic matrices of Herculex I -fed larvae and isoline-fed larvae indicated that Herculex I did not result in damage to the peritrophic matrix that could facilitate subsequent baculovirus infection. Additional research is required to further delineate the physiological basis for enhanced baculovirus infection following exposure to sublethal doses of Bt toxins.