Abstract
Two kinds of neutral protease activities in lymph nodes from Lewis rats with acute experimental allergic encephalomyelitis (EAE) have been separated and partially purified and characterized. A soluble enzyme preparation enriched by gel filtration and ion-exchange chromatography hydrolyzes myelin basic protein, polylysine, and other basic proteins with an optimum pH at 6.0–6.5. It is inhibited byp-chloromercuribenzoate, and thus appears to be a mixture of thiol proteases. Another fraction containing proteolytic enzyme activity is strongly bound to the insoluble lymph node residue, and it also hydrolyzes myelin basic protein and histone, but not polylysine. It has a pH optimum above 7.5, is inhibited by phenylmethylsulfonyl fluoride, thus resembling elastase, but does not hydrolyze elastin-Congo red. The insoluble enzyme preparation hydrolyzes basic protein to 4–5 peptides in a pattern on polyacrylamide gels resembling that of the hydrolysis of basic protein by whole lymphocytes; the soluble enzyme mixture produces small fragments not retained on gels. Lymphocytes are a major component of the cells inflitrating the nervous system in experimental allergic encephalomyelitis, and neutral proteases contained in these cells may contribute to the degradation of myelin, especially of the basic protein.
Similar content being viewed by others
References
Smith, M. E. 1978. Basic protein hydrolysis in lymphocytes of Lewis rats with experimental allergic encephalomyelitis. Adv. Exp. Med. Biol. 100:347–363.
Smith, M. E. 1979. Neutral protease activity in lymphocytes of Lewis rats with acute experimental allergic encephalomyelitis. Neurochem. Res. 4:689–702.
Smith, M. E. 1976. A lymph node neutral proteinase acting on myelin basic protein. J. Neurochem. 27:1077–1082.
Smith, M. E. 1968. The turnover of myelin in the adult rat. Biochim. Biophys. Acta 164:285–293.
Moore, S., andStein, W. H. 1954. A modified ninhydrin reagent for the photometric determination of amino acids and related compounds. J. Biol. Chem. 211:907–913.
Lowry, O. H., Rosebrough, N. J., Farr, A. L., andRandall, R. J. 1951. Protein measurement with the Folin phenol reagent. J. Biol. Chem. 193:265–275.
Weber, K., andOsborne, M. 1969. The reliability of molecular weight determinations by dodecylsulfate-polyacrylamide gel electrophoresis. J. Biol. Chem. 244:4406–4412.
Gray, W. R. 1972. End-group analysis using dansyl chloride. Pages 121–138,in Hirs, C. H. W., andTimasheff, S. N. (eds.), Methods in Enzymology, Vol. XXV, Enzyme Structure, Part B, Academic Press, New York.
Neuhoff, V. 1973. Microdetermination with dansyl chloride. Pages 85–147,in Micromethods of Molecular Biology, Springer, New York.
Woods, K. R., andWang, K. T. 1967. Separation of dansyl-amino acids by polyamide layer chromatography. Biochim. Biophys. Acta 133:369–370.
Yago, N., andBowers, W. E. 1975. Unique cathepsin-D-type proteases in rat thoracic duct lymphocytes and in rat lymphoid tissues. J. Biol. Chem. 250:4749–4754.
Grayzel, A. I., Hatcher, V. G., andLazarus, G. S. 1975. Protease activity of normal and PHA stimulated human lymphocytes. Cell. Immunol. 18:210–219.
Hatcher, V. B., Oberman, M. S., Lazarus, G. S., andGrayzel, A. I. 1978. A cytotoxic proteinase isolated from human lymphocytes. J. Immunol. 120:665–670.
Roboz-Einstein, E., Csejtey, J., Davis, W. J., Lajtha, A., andMarks, N. 1969. Enzymatic degradation of the encephalitogenic protein. Add. ad Int. Arch. Allergy 36:363–375.
Hallpike, J. F., andAdams, C. W. M. 1969. Proteolysis and myelin breakdown: A review of recent histochemical and biochemical studies. Histochem. J. 1:559–578.
Smith, M. E. 1977. The role of proteolytic enzymes in demyelination in experimental allergic encephalomyelitis. Neurochem. Res. 2:233–246.
Cammer, W., Bloom, B. R., Norton, W. T., andGordon, S. 1978. Degradation of basic protein in myelin by neutral proteases secreted by stimulated macrophages. A possible mechanism of inflammatory demyelination. Proc. Natl. Acad. Sci. U.S.A. 75:1554–1558
Barrett, A. J. 1977. Cathepsin B and other thiol proteinases. Pages 181–208,in Barrett, A. J. (ed.), Proteinases in Mammalian Cells and Tissues. North Holland Publishing Co., New York.
Starkey, P. M. 1977. Elastase and cathepsin G: The serine proteinases of human neutrophill leucocytes and spleen. Pages 57–89,in Barrett, A. J. (ed.), Proteinases in Mammalian Cells and Tissues, North Holland Publishing Co., New York.
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Smith, M.E., Chow, S.H. & Rolph, R.H. Partial purification and characterization of neutral proteases in lymph nodes of rats with experimental allergic encephalomyelitis. Neurochem Res 6, 901–912 (1981). https://doi.org/10.1007/BF00965048
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF00965048