Photochemically Modified α-Crystallin: A Model System for Aging in the Primate Lens¶

Abstract

The purpose of this study was to quantitatively study the changes that occur upon irradiation of 3-hydroxykynurenine (3-HK) in the presence of ␣-crystallin under conditions similar to those in the lens. The samples were prepared in 10 mM phosphate buffer at pH 7.4, bubbled with O 2 or Ar and irradiated with 300-400 nm light. The amount of light absorbed by the samples (I abs ) was measured using azobenzene as an actinometer. Modifications to ␣-crystallin were monitored by ultraviolet-visible and fluorescence spectroscopy. Aerobic samples had increased absorption around 320 nm and above 400 nm while the 3-HK maximum at 368 nm decreased. The isolated modified protein showed that there was increased absorption throughout the spectrum. Changes in the anaerobic samples were similar to those of the aerobic but occurred more slowly. As irradiation time increased fluorescence emission of the isolated protein red shifted and quantum yields of fluorescence (⌽ f ) were calculated at different irradiation time intervals by comparison to 3-HK. By comparing OD 320 /OD 365 for the model system to values from primate lenses, I abs can be correlated with age and transmission of the sample in the blue region of the spectrum and thus allows lenticular aging to be quantitated.

Time-resolved and Steady-state Fluorescence Spectroscopic Studies of the Human Lens with Comparison to Argpyrimidine, Pentosidine and 3-OH-kynurenine¶

“…Thus, its fluorescence contribution would be negligible even if 3‐OH‐kynurenine contributes to the yellow hue of the human lens. It would be of interest to compare argpyrimidine and pentosidine to various derivatives of kynurenine to lens proteins, which are believed to be much more fluorescent than kynurenine itself (12,17,25). However, to our knowledge, those compounds are not commercially available.…”

Monitoring the Accumulation of Lipofuscin in Aging Murine Eyes by Fluorescence Spectroscopy

“…The goal of this study was to monitor the accumulation of lipofuscin in murine eyes, to compare these spectra with those from eyes from other species, and to compare these data with other tissues. Previous attempts to characterize age‐related changes in eyes were based on absorption study of primate lens (34,35) and transmission of light to the human retina (36). Roberts et al .…”

Non-enzymatic glycation of α-crystallin as an in vitro model for aging, diabetes and degenerative diseases

“…In α-crystallin, tryptophans are not located at the N terminus and hence do not have a free amino group to participate in the Maillard reaction. Dillon et al have studied extensively on the photo-degradation products of alpha crystallin and showed that they were primarily from tryptophan oxidation (Dillon 1991;Ervin et al 2001;Finley et al 1997Finley et al , 1998Gaillard et al 2000;McDermott et al 1991;Roberts et al 1991).…”