EurekaMag.com logo
+ Site Statistics
References:
53,517,315
Abstracts:
29,339,501
+ Search Articles
+ Subscribe to Site Feeds
EurekaMag Most Shared ContentMost Shared
EurekaMag PDF Full Text ContentPDF Full Text
+ PDF Full Text
Request PDF Full TextRequest PDF Full Text
+ Follow Us
Follow on FacebookFollow on Facebook
Follow on TwitterFollow on Twitter
Follow on Google+Follow on Google+
Follow on LinkedInFollow on LinkedIn

+ Translate

The identification of the major excreted protein (MEP) from a transformed mouse fibroblast cell line as a catalytically active precursor form of cathepsin L



The identification of the major excreted protein (MEP) from a transformed mouse fibroblast cell line as a catalytically active precursor form of cathepsin L



Biochemical Journal 248(2): 449-454



The major excreted protein (MEP) purified from Kirsten-virus-transformed 3T3 fibroblasts and mature human cathepsin L were compared in respect to a number of catalytic criteria and found to be similar. The Mr of MEP is 39,000, whereas that of mature human cathepsin L is 30,000. Sequence data suggested that MEP could be a pro-form of mouse cathepsin L. Both enzymes acted on the synthetic substrate benzyloxycarbonyl-Phe-Arg-7-(4-methyl)coumarylamide with similar catalytic constants and acted optimally at pH 5.5. Both were rapidly inactivated by the active-site-directed inhibitors benzyloxycarbonyl-Phe-Phe-diazomethane and L-3-carboxy-trans-2,3-epoxypropionyl-leucylamido-(4-guanidin o)butane, and furthermore, 3H-labelled L-3-carboxy-trans-2,3-epoxypropionyl-leucylamido-(4-acetamid o)butane, which binds covalently to the heavy chain of mature cathepsin L, also bound to MEP. MEP autolyses rapidly at pH 3.0 to give lower-Mr (35,000 and 30,000) forms, but all forms react with the radiolabelled inhibitor. No autolysis occurred above pH 5.0. MEP hydrolysed azocasein at pH 5.0, demonstrating that it is capable of hydrolysing protein substrates without autolytic activation. Unlike mature forms of cathepsin L, MEP is stable, but not active, at neutral pH. The present work shows that cathepsin L can be secreted as a higher-Mr precursor that is stable in extracellular fluids but only active where local pH values fall below 6.0. These results suggest that the extra N-terminal peptide on MEP is not an activation peptide, but is a regulatory peptide affecting the pH-stability and activity of mouse cathepsin L.

(PDF same-day service: $19.90)

Accession: 006689563

Download citation: RISBibTeXText

PMID: 3435459

DOI: 10.1042/bj2480449



Related references

The major excreted protein of transformed mouse cells and cathepsin l have similar protease specificity. Biochemical & Biophysical Research Communications 139(1): 156-162, 1986

The major excreted protein (MEP) of transformed mouse cells and cathepsin L have similar protease specificity. Biochemical and Biophysical Research Communications 139(1): 156-162, 1986

The preparation of catalytically active human cathepsin B from its precursor expressed in Escherichia coli in the form of inclusion bodies. European Journal of Biochemistry 229(2): 533-539, 1995

The major excreted protein of transformed mouse fibroblasts localization and regulation. Journal of Cell Biology 95(2 PART 2): 390A, 1982

Hybridization of a mouse T-cell lymphocyte line (HB1) with a polyoma virus-transformed mouse fibroblast line. Experimental Cell Research 134(2): 445-456, 1981

A catalytically active high-Mr form of human cathepsin B from sputum. Biochemical Journal 254(3): 693-699, 1988

Expression of eosinophil granule major basic protein and its precursor form, proMBP, by the human promyelocytic leukemia cell line. Journal of Allergy & Clinical Immunology 93(1 PART 2): 195, 1994

Cloning and expression of the gene for the major excreted protein of transformed mouse fibroblasts. A secreted lysosomal protease regulated by transformation. Journal of Biological Chemistry 263(1): 254-261, 1988

Effect of tumor necrosis factor-alpha on a cell line transformed by a secreted form of human fibroblast growth factor-1 gene and on its parental cell line. Cancer Letters. 89(1): 49-54, 1995

A catalytically active high molecular weight form of human cathepsin b from sputum. Biochemical Journal 254(3): 693-700, 1988

The major excreted protein of transformed tumor promoter and platelet derived growth factor treated mouse fibroblasts is an acid activated protease. Journal of Cell Biology 101(5 PART 2): 55A, 1985

Isolation of an oncogene from a mouse fibroblast cell line transformed with 5 aza 2 deoxycytidine. Proceedings of the American Association for Cancer Research Annual Meeting 32: 134, 1991

Restoration of normal growth control to a transformed mouse fibroblast cell line induced by differentiation agents. Federation Proceedings 43(3): ABSTRACT 2881, 1984

Retinoic acid restores normal growth control to a transformed mouse embryo fibroblast cell line. Cancer Letters 33(1): 33-44, 1986