Evidence for the existence of both proteasomes and a novel high molecular weight peptidase in Entamoeba histolytica

Autor(en): Scholze, H
Frey, S
Cejka, Z
BakkerGrunwald, T
Stichwörter: Biochemistry & Molecular Biology; COMPLEXES; PROTEOLYSIS; PURIFICATION
Erscheinungsdatum: 1996
Herausgeber: AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC
Journal: JOURNAL OF BIOLOGICAL CHEMISTRY
Volumen: 271
Ausgabe: 11
Startseite: 6212
Seitenende: 6216
Zusammenfassung: 
To screen for high molecular weight proteases in Entamoeba histolytica, we subjected a soluble amebal extract to density gradient centrifugation and tested the fractions for activity against the chymotryptic peptide substrate, Suc-leucyl-leucyl-valyl-tyrosyl-4-methylcoumaryl-7-amide. Two peaks of activity, of approximately 11 and 20 S, were clearly separated. Based on SDS-electrophoretic pattern and immunoblot analysis, we ascribe the 20 S activity to proteasomes. The 11 S protein was purified from amebal homogenates by a series of chromatographic steps. As determined by molecular sieve chromatography and nondenaturing gel electrophoresis, the native complex had an apparent M(r) of 385,000 /- 10%. On SDS gels, the purified enzyme exhib ited a single band of M(r) 62,000 that yielded a single N-terminal sequence, indicating that the preparation was homogeneous and that the native complex consisted of six very similar or identical subunits. The enzyme preferred peptides with aromatic residues at the P-1 position and had low but distinct activity toward azocasein. We conclude that the 11 S enzyme is a novel high molecular weight protease that is distinct from proteasomes.
ISSN: 00219258
DOI: 10.1074/jbc.271.11.6212

Show full item record

Google ScholarTM

Check

Altmetric