Degraded myelin-associated glycoprotein (dMAG) formation from pure human brain myelin-associated glycoprotein (MAG) is not mediated by calpain or cathepsin L-like activities.
The myelin-associated glycoprotein (MAG) is a transmembrane cell adhesion molecule participating in myelin formation and maintenance. Calcium-activated/-dependent proteolysis of myelin-associated glycoprotein by calpain and cathepsin L-like activities has already been detected in purified myelin fractions, producing a soluble fragment, called degraded (d)MAG, characterized by the loss of the transmembrane and cytoplasmic domains. Here, we demonstrate and analyze dMAG formation from pure human brain myelin-associated glycoprotein. The activity never exhibited the high rate previously reported in human myelin fractions. Degradation is time-, temperature-, buffer- and structure-dependent, is inhibited at 4 degrees C and by denaturation of the sample, and is mediated by a trans-acting factor. There is no strict pH dependency of the proteolysis. Degradation was inhibited by excess aprotinin, but not by 1-10 micro g/mL aprotinin and was not eliminated by the use of an aprotinin-sepharose matrix during the purification. dMAG formation was not enhanced by calcium, nor inhibited by a wide variety of protease inhibitors, including specific calpain and cathepsin L inhibitors. Therefore, while cysteine proteases may be present in human myelin membrane fractions, they are not involved in dMAG formation from highly purified human brain myelin-associated glycoprotein preparations.
Päiväläinen
Satu
S
Department of Pathology, University of Oulu, Oulu, Finland.
Suokas
Marko
M
Lahti
Outi
O
Heape
Anthony M
AM
D000368:Aged; D000369:Aged, 80 and over; D001923:Brain Chemistry; D002021:Buffers; D002118:Calcium / Q000737:chemistry; D002154:Calpain / Q000737:chemistry / Q000378:metabolism*; D056668:Cathepsin L; D002403:Cathepsins / Q000737:chemistry / Q000378:metabolism*; D002614:Chelating Agents / Q000737:chemistry; D003546:Cysteine Endopeptidases; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008875:Middle Aged; D018992:Myelin-Associated Glycoprotein / Q000737:chemistry* / Q000378:metabolism*; D011480:Protease Inhibitors / Q000737:chemistry; D013696:Temperature
D002021:Buffers; D002614:Chelating Agents; D018992:Myelin-Associated Glycoprotein; D011480:Protease Inhibitors; D002403:Cathepsins; D002154:Calpain; D003546:Cysteine Endopeptidases; C534281:CTSL protein, human; D056668:Cathepsin L; D002118:Calcium