Physical and chemical changes in alpha-crystallin during maturation of lens fibers

• Published in: Experimental eye research Vol. 18; no. 4; pp. 383 - 393
• Main Author: Li, Lu-Ku
• Format: Journal Article
• Language: English
• Published: England Elsevier Ltd 01.04.1974
• Subjects: Amino Acids - analysis; Animals; Cattle; Chromatography, DEAE-Cellulose; Circular Dichroism; Crystallins - analysis; Electrophoresis, Polyacrylamide Gel; Lens, Crystalline - growth & development; Molecular Weight; Protein Denaturation; Spectrum Analysis; Ultracentrifugation
• ISSN: 0014-4835; 1096-0007
• DOI: 10.1016/0014-4835(74)90116-X

A mechanical separation of calf lens into three fractions composed of outer cortex, inner cortex and nucleus provided material representing three phases of fiber maturation. Separation of the lenticular proteins in each fraction was achieved by gel filtration on agarose 15 m. The amount of high mol. wt. alpha-crystallin aggregates was found to increase appreciably as the fibers age. The amount of low mol. wt. alpha-crystallin, however, decreases while the average size gradually increases as the fibers are displaced toward the center of the lens. An increase in sedimentation coefficient corresponding to an increase of 50% in mol. wt. was found when the low mol. wt. alpha-crystallin from the outer cortex was compared to that from the nuclear region. No detectable change in the amino acid or the subunit composition accompanied this increase in size. The circular dichroism (CD) of these crystallins also revealed no conformation changes of the polypeptide backbone. Unique to the CD spectrum for the nuclear alpha-crystallin are minima at 235, 214 and 206 nm. This observation was corroborated by far u.v. spectroscopy to 195 nm. The loss of these CD minima as well as changes in the u.v. profile that can be effected by 7 m-urea suggests the presence of chromophore(s) attached non-covalently to the nuclear protein. One or more other chromophore(s) common to all low mol. wt. alpha-crystallins were revealed by a 40% reduction in the extinction at 195 nm after urea treatment of the cortical alpha-crystallins with no associated change in the CD spectrum. The 20% reduction in extinction for the nuclear protein suggests the presence of similar chromophore(s). Deaggregation followed by reaggregation not only markedly diminished the size of the low mol. wt. alpha-crystallins but also drastically reduced the disparities in size of this protein among the three fiber fractions. These results suggest that chromophore(s) removed by urea treatment may play a decisive role in determining the size of the alpha-crystallin marcromolecule during the maturation of the lens fibers.