Cleavage of beta crystallins during maturation of bovine lens.
OBJECTIVE
(1) Identify major crystallin proteins in fetal and adult bovine lens, (2) examine the N-termini of beta-crystallins for truncation, and (3) determine if the protease m-calpain (EC 3.4.22.17) is responsible for the cleavage of bovine beta-crystallins during maturation.


METHODS
Crystallins from fetal and adult bovine lenses were analyzed by one and two-dimensional electrophoresis and Edman sequencing of separated proteins and their tryptic fragments. Identical techniques were used to analyze crystallins following their incubation with purified m-calpain.


RESULTS
The identities of the major crystallins and several additional crystallin species missing portions of their N-terminal extensions were identified in the fetal bovine lens. Besides the previously identified form of betaB1 missing 15 residues from its N-terminus, forms of betaA3 >missing 11 and 22 residues were identified. With aging, the betaA3 (-22) species became a major protein in the adult bovine lens, and minor forms of betaB2 and betaB3 missing 8 and 22 residues from their N-termini, respectively, appeared. Purified m-calpain cleaved within the N-terminal extensions of bovine beta-crystallins and removed: 12 or 15 residues from betaB1; 8 residues from betaB2; 5 or 10 residues from betaB3; and 11 or 17 residues from betaA3.


CONCLUSIONS
Based on the cleavage sites in vitro, m-calpain may be partially responsible for cleavage of bovine betaB1, betaB2, and betaA3 during lens maturation. However, the preference of m-calpain to remove 12 residues from betaB1, and 11 and 17 residues from betaA3, suggested that the betaB1 (-15) and betaA3 (-22) species found in vivo were produced by a different protease. This unidentified protease may have a preference for the asparagine-proline-X-proline sequence found in the N-terminal extensions of betaB1 and betaA3.
Shih
M
M
Department of Oral Molecular Biology, Oregon Health Sciences University, Portland, OR 97201, USA.
Lampi
K J
KJ
Shearer
T R
TR
David
L L
LL
D000375:Aging; D000595:Amino Acid Sequence; D000818:Animals; D002154:Calpain / Q000378:metabolism; D002417:Cattle; D003459:Crystallins / Q000737:chemistry* / Q000378:metabolism*; D007908:Lens, Crystalline / Q000737:chemistry* / Q000196:embryology / Q000201:enzymology / Q000254:growth & development; D008969:Molecular Sequence Data; D017421:Sequence Analysis
D003459:Crystallins; D002154:Calpain