Extraction, purification and characterization of the crystallin protein of cataractous eye lens nucleus.
The purpose of this study is to separate and identify the crystallin protein present in the nucleus of a human cataractous eye lens. Cataractous lenses were collected from different eye hospitals from patients of different etiologies with ages between 40 and 80 years. Lens nucleus proteins were extracted into four fractions on the basis of their solubility in different media by applying a reported method. These fractions were buffer-soluble proteins (PS), urea-soluble proteins (PU), yellow fraction proteins (PY) and insoluble proteins (PI). All three soluble fractions were subjected to HPLC and GPC analysis. Both HPLC and GPC analysis showed that each fraction contains α-, β- and γ-crystallins, a major class of protein present in the lenses of vertebrates. Various chromatographic parameters including precision, accuracy and linearity have been evaluated. Studies of water-insoluble crystallins using sodium dodecylsulphate polyacrylamide gel electrophoresis (SDS-PAGE) have demonstrated extreme homogeneity with evidence of major components with molecular masses of 18-70 kDa, similar to the crystallin of the water-soluble portion. The method was found to be suitable for the analysis of various isomers of crystallin protein present in human cataractous eye lens nuclei. The detailed results of the GPC are discussed. This study provides the first HPLC and GPC analysis of a human cataractous eye lens nucleus.
Department of Chemistry, University of Sargodha, Sargodha 40100, Pakistan. msherawan@yahoo.com.
Muhammad
M
Sher
Ayesha
A
Hameed
Sobia
S
Noreen
Muhammad
M
Fayyaz-ur-Rehman
Muhammad Ajaz
MA
Hussain
Syed Nasir Abbas
SN
Bukhari
D005591:Chemical Fractionation / Q000379:methods*; D002850:Chromatography, Gel; D003459:Crystallins / Q000737:chemistry* / Q000302:isolation & purification*; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D007907:Lens Nucleus, Crystalline / Q000737:chemistry*; D012995:Solubility
D003459:Crystallins