title: Differential influence of proteolysis by calpain 2 and Lp82 on in vitro precipitation of mouse lens crystallins.
abstract: The purpose of the present study was to compare the susceptibility of crystallins proteolyzed by ubiquitous calpain 2 and by lens-specific calpain Lp82 to insolubilization. To test this, transgenic (TG) mice expressing a calpain 2, in which the active site cysteine 105 was mutated to alanine, were produced. Expression of mutated calpain 2 was driven in lens by coupling the mutated gene to the betaB1-crystallin promoter. Light scattering was measured in solutions of lens proteins after activation of endogenous calpain 2 and/or Lp82. Mass spectrometric analysis was performed to determine the cleavage sites and the calpain responsible for insolubilization of crystallins. Lens proteins from TG mice incubated in vitro with calcium showed higher light scattering compared to proteins from wild type (WT) mice. alphaA-crystallin from TG mice was proteolyzed by Lp82. In contrast, alphaA-crystallin in lenses from WT mice were proteolyzed by both calpain 2 and Lp82. These results suggested that Lp82-induced proteolysis of crystallins caused increased susceptibility of truncated crystallins to in vitro precipitation. Since Lp82 is highest in young animals, Lp82-induced proteolysis and precipitation may be one of the factors responsible for the cataract formation in young rodents.
authors.affiliation: Research Laboratory, Senju Pharmaceutical Corporation Limited, Kobe 651-2241, Japan. mitsuyoshi-azuma@senju.co.jp
authors.forename: Mitsuyoshi
authors.initials: M
authors.lastname: Azuma
authors.forename: Yoshiyuki
authors.initials: Y
authors.lastname: Tamada
authors.forename: Sayaka
authors.initials: S
authors.lastname: Kanaami
authors.forename: Emi
authors.initials: E
authors.lastname: Nakajima
authors.forename: Yoshikuni
authors.initials: Y
authors.lastname: Nakamura
authors.forename: Chiho
authors.initials: C
authors.lastname: Fukiage
authors.forename: Neil E
authors.initials: NE
authors.lastname: Forsberg
authors.forename: Melinda K
authors.initials: MK
authors.lastname: Duncan
authors.forename: Thomas R
authors.initials: TR
authors.lastname: Shearer
mesh_terms: D000818:Animals; D002154:Calpain / Q000235:genetics / Q000378:metabolism*; D011232:Chemical Precipitation; D003459:Crystallins / Q000737:chemistry / Q000378:metabolism*; D008027:Light; D051379:Mice; D008822:Mice, Transgenic; D009154:Mutation
chemical_list: D003459:Crystallins; D002154:Calpain; C118624:calpain Lp82