Characterization and regulation of lens-specific calpain Lp82

Chiho Fukiage, Emi Nakajima, Ma Hong, Mitsuyoshi Azuma, Thomas R. Shearer

Research output: Contribution to journalArticle

33 Scopus citations

Abstract

Eye tissues contain splice variants of muscle-preferred p94 (calpain 3), such as lens-specific Lp82 and Lp85, retina-specific Rt88, and cornea-specific Cn94. The purpose of the present experiment was to analyze the activation and regulation of the best characterized p94 splice variant, Lp82. Recombinant rat Lp82 (rLp82) was expressed using the baculovirus system, purified with Ni-NTA affinity and DEAE-ion exchange chromatographies, and characterized by SDS-PAGE, casein zymography, and immunoblotting. After incubation with calcium, rLp82 autolyzed into two major fragments at ∼60 and 22 kDa. Sequencing of the autolytic fragments showed loss of three amino acids from the N terminus and cleavage near the IS2 region. Also, Lp82 and calpain 2 were found to hydrolyze each other. Calpastatin inhibited calpain 2 activity, but not Lp82. Homology modeling suggested that the lack of inhibition of Lp82 by calpastatin was due to molecular clashes at the unique AX1 region of Lp82. Lp82 also hydrolyzed calpastatin. These results suggested that Lp82 might reg. ulate other calpain activities and cause hydrolysis of substrates such as crystallins during lens cataract formation.

Original languageEnglish (US)
Pages (from-to)20678-20685
Number of pages8
JournalJournal of Biological Chemistry
Volume277
Issue number23
DOIs
StatePublished - Jun 7 2002

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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