title: BetaB2-crystallin undergoes extensive truncation during aging in human lenses.
abstract: Based on the present literature, it is unclear whether betaB2-crystallin undergoes age-related truncation in human lenses. To answer this question, the purpose of this study was to determine in vivo truncation of betaB2-crystallin in human lenses during aging by examining its fragments in the beta(H)-crystallin fraction. The WS-protein fraction was isolated from lenses of desired ages and separated by a size-exclusion Agarose A 1.5m column to recover alpha-, beta(H)-, beta(L)-, and gamma-crystallin fractions. The beta(H)-crystallin fractions, isolated from lenses of 24- and 70-year-old donors, were utilized for two-dimensional (2D) gel electrophoresis (isoelectric focusing in the first dimension followed by SDS-PAGE in the second dimension). The partial N-terminal sequences of the desired fragments (Molecular weights [M(r)]<18-19kDa) from a 2D-gel of WS-proteins from lenses of a 70-year-old donor were determined. More than 37 crystallin fragments with M(r) between 4 and 19kDa were observed on a 2D-gel. Nine fragments in beta(H)-crystallin fraction were from betaB2-crystallin but additional single fragments of alphaA-, gammas-, betaA4, and of either gammaB-, gammaC- or gammaD-crystallins were also observed. Seven cleavage sites in the betaB2-crystallin were identified, which included two sites at Q(7)-A(8) and A(8)-G(9) bonds in the N-terminal extension, two sites at E(46)-K(47) and G(49)-S(50) bonds in the motif 1, one site at S(94) -S(95) in the motif 2, and two sites at N(115)-F(116) and Q(135)-Y(136) in motif 3. No fragments with cleavage in the motif 4 and C-terminal extension of betaB2-crystallin were seen. Apparently, three betaB2-crystallin fragments with only N-terminal cleavage and five with both N- and C-terminal cleavages were observed. Additional fragments with cleavage sites at Q(54)-Y(55) in alphaA-crystallin, at E(112)-N(113) in betaA4-crystallin, at G(4)-T(5) in gammas-crystallin, at M(69)-G(70) in either gammaB-, gammaC- or gammaD-crystallins (three have identical sequences at the cleaved bond), and at G(1)-K(2) in gammaB or gammaC (both have identical sequences at the cleavage site) were observed.Conclusions. The results showed that betaB2-crystallin undergoes age-related truncation producing fragments with M(r) between 4 and 19kDa that existed in the beta(H)-crystallin oligomer. The beta(H)-crystallin fraction also contained single fragments of alpha-, betaA4-, gammas-, and other gamma-crystallins.
authors.affiliation: Department of Physiological Optics, School of Optometry, University of Alabama at Birmingham, Worrell Building, 924 South 18th Street, Birmingham, AL 35294-4390, USA. Srivastava@icare.pt.opt.uab.edu
authors.forename: O P
authors.initials: OP
authors.lastname: Srivastava
authors.forename: K
authors.initials: K
authors.lastname: Srivastava
mesh_terms: D000328:Adult; D000368:Aged; D000375:Aging / Q000502:physiology*; D000595:Amino Acid Sequence; D015180:Electrophoresis, Gel, Two-Dimensional; D006801:Humans; D007908:Lens, Crystalline / Q000737:chemistry / Q000378:metabolism*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D038206:beta-Crystallin B Chain / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism*
chemical_list: D010446:Peptide Fragments; D038206:beta-Crystallin B Chain; C113634:beta-crystallin B2