This study is to characterize the proteins of the human lens with development, aging, and cataractogenesis. The lens consists of a few very high abundance proteins called the crystallins and several hundred lesser abundance proteins, including the metabolic enzymes and cytoskeletal proteins. During the life of the lens, the proteins become extensively modified. It has been thought that these modifications are involved in cataract formation, but the results of this study have demonstrated that the major crystallin modifications are developmentally regulated. Methodologies have been developed to yield good separation of these proteins by using two-dimensional polyacrylamide gel electrophoresis. Due to the extensive modifications, identification of the proteins is based on immunotechniques, sequencing, and mass spectrometry. Normal donor lenses varying in age from fetal to 70 years were analyzed. Lenses were dissected by peeling the shells of fiber cells. This method yields defined populations of fiber cells from the cortex and each of the developmentally defined nuclear regions. The protein patterns in each of the cortical regions are distinguishable as cortex with the characteristic large protein spots corresponding to each of the major crystallins. The protein patterns of the nuclear regions are all similar to each other but clearly unique from that of the cortical regions. Only protein species that migrate in new positions are present. These include more acidic forms and low molecular weight fragments of the crystallins. Crystallin modifications in the nuclear regions begin to occur shortly after birth and are not related to cataractogenesis. Results of studies on the water-soluble crystallins demonstrate that the crystallin modifications start to occur in the second layer of the cortex in normal lenses of all ages. There appears to be no preferential insolubilization of the modified crystallins. The proteins throughout the lens are being identified, yielding a database of information on the normal human lens. The protein patterns of numerous cataracts of different etiologies have been determined. The proteins of the nuclear regions of nuclear cataracts are less soluble and contain increased levels of disulfide bonds compared with the nuclear regions of normal lenses, but the protein patterns of the nuclear regions of nuclear cataracts are similar to those from normal age-matched lenses. In more advanced nuclear cataracts, there is a preferential decrease in the solubility of modified forms of one of the crystallins. Further studies on the solubility of the crystallins are in progress. Lens proteins from donors with trisomies and syndromes such as Down's are being analyzed.
