title: Sequential limited proteolysis of myelin basic protein by neutral protease activities of bovine brain.
abstract: Acid extracts of delipidated white matter of bovine brain were prepared, and their proteolytic activities toward myelin basic protein (MBP) were evaluated at pH 3 and pH 7. This was done by measuring changes in total protein using a selective dye-binding assay, and by evaluating peptide patterns by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and densitometry. At pH 7 greater than 50% of total protein and about 75% of MBP were degraded after 48 h, whereas at pH 3 it was less than 20% altogether. Neutral proteolysis of MBP entailed up to 12 different proteolytic peptide fragments in the molecular weight range of 17.5 to 6 kd. Its enzymatic nature was verified using protease inhibitors, including N-ethylmaleimide, phenylmethylsulfonyl fluoride, o-phenanthroline, and EDTA, as well as pepstatin A and alpha 2-macroglobulin. Both transient changes in percentages of some intermediate peptides and differential effects of individual inhibitors on electrophoretic peptide patterns strongly suggest a sequential type of limited proteolysis. The results also indicate that acid extracts contained several endopeptidases of which a cysteine protease appears to initiate the breakdown of MBP.
authors.forename: H H
authors.initials: HH
authors.lastname: Berlet
authors.forename: H
authors.initials: H
authors.lastname: Ilzenhöfer
mesh_terms: D000818:Animals; D001921:Brain / Q000201:enzymology*; D002417:Cattle; D004492:Edetic Acid / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D010450:Endopeptidases / Q000378:metabolism*; D005033:Ethylmaleimide / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008970:Molecular Weight; D004676:Myelin Basic Protein / Q000378:metabolism*; D015260:Neprilysin; D010436:Pepstatins / Q000494:pharmacology; D010446:Peptide Fragments / Q000378:metabolism; D010618:Phenanthrolines / Q000494:pharmacology; D010664:Phenylmethylsulfonyl Fluoride / Q000494:pharmacology; D011480:Protease Inhibitors / Q000494:pharmacology; D000511:alpha-Macroglobulins / Q000494:pharmacology
chemical_list: D004676:Myelin Basic Protein; D010436:Pepstatins; D010446:Peptide Fragments; D010618:Phenanthrolines; D011480:Protease Inhibitors; D000511:alpha-Macroglobulins; C014195:Streptomyces pepsin inhibitor; D010664:Phenylmethylsulfonyl Fluoride; D004492:Edetic Acid; D010450:Endopeptidases; D015260:Neprilysin; D005033:Ethylmaleimide; C031375:pepstatin; C025205:1,10-phenanthroline