Lp82 is the Dominant Form of Calpain in Young Mouse Lens

• Published in: Experimental eye research Vol. 68; no. 4; pp. 447 - 456
• Main Authors: MA, H., HATA, I., SHIH, M., FUKIAGE, C., NAKAMURA, Y., AZUMA, M., SHEARER, T.R.
• Format: Journal Article
• Language: English
• Published: London Elsevier Ltd 01.04.1999; Elsevier
• Subjects: Amino Acid Sequence; Animals; Base Sequence; Biological and medical sciences; Calpain; casein zymography; Chromatography, High Pressure Liquid; Cloning, Molecular; Crystallins - metabolism; Cysteine Endopeptidases - genetics; Cysteine Endopeptidases - isolation & purification; Cysteine Endopeptidases - metabolism; Electrophoresis, Polyacrylamide Gel; Enzyme-Linked Immunosorbent Assay; Eye and associated structures. Visual pathways and centers. Vision; Female; Fundamental and applied biological sciences. Psychology; immunoblot; Immunoblotting; Isoenzymes - genetics; Isoenzymes - isolation & purification; lens maturation; Lens, Crystalline - enzymology; Lp82; m-calpain; Mice; Mice, Inbred Strains; molecular cloning; Molecular Sequence Data; mouse lens; mRNA; Rats; Rats, Sprague-Dawley; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger - analysis; Sequence Homology, Amino Acid; Sequence Homology, Nucleic Acid; Vertebrates: nervous system and sense organs; casein zymography; lens maturation; immunoblot; molecular cloning; m-calpain; Lp82; mRNA; mouse lens; Molecular form; Enzyme; Isozyme; Cysteine endopeptidases; Rodentia; Calpain; Eye; Visual system; Peptidases; Vertebrata; Mammalia; Mouse; Hydrolases; Lens; Molecular cloning
• ISSN: 0014-4835; 1096-0007
• DOI: 10.1006/exer.1998.0625

The purpose of this study was to characterize Lp82 calpain in normal mouse. Lp82 is a lens-specific, calcium-activated isozyme from the calpain super family of cysteine proteases (EC 34.22.17). RT-PCR and molecular cloning were performed on total RNA from 12 day-old mice. Lp82 and m-calpain protein levels and proteolytic activities in lenses were measured by casein zymography, immunoblotting, and ELISA after partial purification by DEAE-HPLC. The 2334-bp cDNA encoding for mouse Lp82 contained a single large open reading frame encoding a protein of 709 amino acid residues with a calculated molecular weight of 82.2 kDa and a predicted pI of 5.8. The amino acid sequence of mouse lens Lp82 was 99% homologous to rat lens Lp82. As in rat, mouse lens Lp82 showed a uniqueN-terminus and deletion of the IS1 and IS2 regions. In contrast to rat, Lp82 was the dominant calpain in young mouse lens. Lp82 was lens-specific, and the lens nucleus contained the highest specific activity of Lp82 and very little m-calpain. Endogenous Lp82 in lens soluble proteins was activated by addition of calcium and caused limited proteolysis of crystallins even in the presence of large amounts of recombinant domain I from the natural calpain inhibitor calpastatin. Loss of Lp82 protein accompanied aging of mouse lens. Lp82 may be responsible for a major portion of crystallin proteolysis occurring during normal lens development and maturation, or during cataract formation in young mice.