Red edge excitation shifts of crystallins and intact lenses: a study of segmental mobility and inter-protein interactions

Abstract

The shift that occurs in the fluorescence emission wavelength upon changing the excitation wavelength towards the red edge of the absorption band is termed red edge excitation shift (REES). We have monitored the REES of intrinsic protein fluorescence of freshly isolated intact lenses, of individual crystallins in their native, denatured and photodamaged states and also of crystallin mixtures. The observed REES values for the lenses from different species are different suggesting that the mobilities and packing of the crystallins may vary with the species. Lens photodamage in all the cases resulted in an increase of REES. Denaturation of crystallins in solution reduces REES and renaturation restores it. Mixtures of α- and β-crystallins prepared either by directly mixing equimolar solutions or mixing them in 4 M urea followed by dialysis (reconstituting) gave similar REES values indicating the absence of any specific interactions in dilute solutions. Possible existence of induced alterations facilitating inter-crystallin interactions at high protein concentration is suggested.