Activation of calpain in lens: A review and proposed mechanism

M. Azuma, C. Fukiage, L. L. David, T. R. Shearer

Research output: Contribution to journalArticle

58 Scopus citations

Abstract

The purpose of these experiments was to develop a hypothesis to explain activation of m-calpain in cataractogenesis observed in rodents. The in vitro model used to study m-calpain activation was to correlate breakdown of the 'reporter' protein α-crystallin with the appearance of activated m-calpain using protein sequencing and casein zymography. Incubation of α-crystallins with m-calpain and Ca2+ caused proteolysis of α-crystallins and accumulation of new polypeptides. E64 and calpain inhibitor each inhibited proteolysis of α-crystallins. The N-terminus of the 80 kDa subunit of m- calpain was blocked at time 0 (pro calpain). After incubation with Ca2+, the remaining 80 kDa subunit of m-calpain gave a N-terminal sequence of KDREAAEGLG, indicating loss of nine amino acid from the N-terminus (autolysed calpain). The new 43 kDa m-calpain fragment also gave a N-terminal sequence of KDREAAEGLC, indicating the same loss of the first nine amino acids on the N-terminus as well as a major loss of the C-terminal half of the subunit (degraded calpain). In contrast, the N-terminus of the 80 kDa subunit of m- calpain remained blocked when E64 was present (unautolysed form). Moreover, the Ca2+ concentration required for proteolysis decreased when calpain was pre-incubated with Ca2+ although proteolysis of α-crystallin required a higher Ca2+ concentration than proteolysis of casein. These data suggested that the sequence of events for m-calpain activation were unantolysed, autolysed and finally degraded calpain. Unautolysed and/or autolysed calpains may be proteolytically active against α-crystallin.

Original languageEnglish (US)
Pages (from-to)529-538
Number of pages10
JournalExperimental Eye Research
Volume64
Issue number4
DOIs
StatePublished - Apr 1997

Keywords

  • Activation
  • Autolysis
  • Calpain
  • Proteolysis
  • Rodents
  • α-crystallin

ASJC Scopus subject areas

  • Ophthalmology
  • Sensory Systems
  • Cellular and Molecular Neuroscience

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