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Cell Biology International (1997) 21, 164174 (Printed in Great Britain)
EXTRA-LYSOSOMAL PROTEOLYSIS AND EXPRESSION OF CALPAINS AND CALPASTATIN IN CULTURED THYROID CELLS
J.‑O. KARLSSON and M. NILSSON
Institute of Anatomy and Cell Biology, Göteborg University, Medicinaregatan 3-5, S-413 90, Göteborg, Sweden
Proteolysis at neutral pH in the soluble fraction of cultured pig thyroid epithelial cells was examined using a synthetic calpain substrate, succinyl-Leu-Tyr-7-amino-4-methylcoumarin. The Ca2+-independent proteolytic activity was largely inhibited by substances known to affect cysteine- and metalloproteases, whereas no or little effects were obtained with inhibitors affecting serine- and aspartic proteases. Addition of Ca2+did not significantly alter the rate of substrate degradation. Biochemical separation via hydrophobic interaction chomatography and Western blotting demonstrated the presence of both m-calpain (40% of total calpain) and μ-calpain (60%) in confluent thyrocytes. Determination of calpastatin activity indicated a 30 times higher level of the inhibitor as compared to total calpain activity. Western blotting showed the presence of a 110
Key words: thyroid, epithelium, proteolysis, calpain, calpastatin, junctions.
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