title: Human micro-calpain: simple isolation from erythrocytes and characterization of autolysis fragments.
abstract: Heterodimeric p-calpain, consisting of the large (80 kDa) and the small (30 kDa) subunit, was isolated and purified from human erythrocytes by a highly reproducible four-step purification procedure. Obtained material is more than 95% pure and has a specific activity of 6-7 mU/mg. Presence of contaminating proteins could not be detected by HPLC and sequence analysis. During storage at -80 degrees C the enzyme remains fully activatable by Ca2+, although the small subunit is partially processed to a 22 kDa fragment. This novel autolysis product of the small subunit starts with the sequence 60RILG and is further processed to the known 18 kDa fragment. Active forms and typical transient and stable autolysis products of the large subunit were identified by protein sequencing. In casein-zymograms only the activatable forms 80 kDa+30 kDa, 80 kDa+22 kDa and 80 kDa+18 kDa displayed caseinolysis.
authors.affiliation: Abteilung für Klinische Chemie und Klinische Biochemie in der Chirurgischen Klinik und Poliklinik-Innenstadt, Klinikum der Ludwig-Maximilians-Universität Munchen, Germany.
authors.forename: D
authors.initials: D
authors.lastname: Gabrijelcic-Geiger
authors.forename: R
authors.initials: R
authors.lastname: Mentele
authors.forename: B
authors.initials: B
authors.lastname: Meisel
authors.forename: H
authors.initials: H
authors.lastname: Hinz
authors.forename: I
authors.initials: I
authors.lastname: Assfalg-Machleidt
authors.forename: W
authors.initials: W
authors.lastname: Machleidt
authors.forename: A
authors.initials: A
authors.lastname: Möller
authors.forename: E A
authors.initials: EA
authors.lastname: Auerswald
mesh_terms: D001329:Autolysis / Q000378:metabolism; D002154:Calpain / Q000097:blood / Q000302:isolation & purification*; D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D004912:Erythrocytes / Q000737:chemistry*; D005470:Fluorometry; D006801:Humans; D020539:Sequence Analysis, Protein
chemical_list: D002154:Calpain