Monitoring Light‐induced Changes in Isolated, Intact Eye Lenses

Abstract

Fluorescence spectral changes occurring upon irradiation with 300 nm light have been monitored in situ in isolated, intact, whole lenses from the eyes of several species. The findings corroborate observations on other individual constituent protein molecules in the solution state, and also reveal features attributable to the supramolecular protein assembly that exists in the whole lens. Irradiation of the lens with 300 nm light causes red shifts in the tryptophan emission spectrum, suggesting alterations in the protein packing in the lens. Intermolecular energy transfer from tryptophan to one of the photoproducts, presumably N‐formylkynurenine (N‐FK), occurs in the condensed‐phase sample. The N‐FK formed is photodegraded efficiently in the lens, indicating that the photodynamic effects of endogenous N‐FK might not be as severe as has been thought. Species variation in the photoevents are evident, particularly in avian lenses that contain the variant δ‐crystallin as the core protein. The photoinduced changes in the near‐UV circular dichroism of δ‐crystallin (which is α‐helical, as opposed to the β‐sheet structure of α‐, β‐, and ‐γ‐crystallins), isolated from chicken lenses, are remarkably different from other crystallins. Irradiation of δ‐crystallin leads to a drastic reduction of circular dichroism intensity in the 250–300 nm region, whereas no changes are seen in the peptide absorption band.

Levels of Reduced Pyridine Nucleotides and Lens Photodamage

“…Tryptophan is the major photochemical target in the lens. The loss of tryptophan fluorescence intensity is taken as a parameter to indicate the lens photodamage (Borkman, 1984;Rao et al, 1987). We have measured tryptophan fluorescence as a function of duration of UV exposure to monitor the lens photodamage.…”

“…Fluorescence spectra were recorded on an AMINCO model SPF 500-C steady state spectrofluorometer. Fluorescence measurements on intact lenses were carried out as described earlier (Rao et al, 1987) using a home built solid state accessory. Emission spectra were obtained upon excitation at 360, 340,280 and 295 nm for each fresh lens.…”

UV-Light Exposed Prion Protein Fails to Form Amyloid Fibrils

“…Other aromatic residues and cysteine also absorb UV light however to a lesser extent. Generation of reactive oxygen species, conformational change and photo-aggregation of γ-crystallin, an eye lens protein, are well documented [29], [30]. In addition to photo-aggregation of γ-crystallin, we have earlier observed conformational dependence of the photo vulnerability of tryptophan residues in different proteins [31].…”

Fluorescence Properties of Isolated Intact Normal Human Corneas

“…Fluorescence measurements of the intact corneas were carried out using a Hitachi F-4010 fluorescence spectrophotometer, with a solidstate accessory in which the sample was mounted and studied, as described earlier (17,18). All spectra were recorded in the corrected spectrum mode with an excitation and emission bandpass of 5 nm each.…”