High level expression and characterisation of Plasmepsin II, an aspartic proteinase from Plasmodium falciparum.
DNA encoding the last 48 residues of the propart and the whole mature sequence of Plasmepsin II was inserted into the T7 dependent vector pET 3a for expression in E. coli. The resultant product was insoluble but accumulated at approximately 20 mg/l of cell culture. Following solubilisation with urea, the zymogen was refolded and, after purification by ion-exchange chromatography, was autoactivated to generate mature Plasmepsin II. The ability of this enzyme to hydrolyse several chromogenic peptide substrates was examined; despite an overall identity of approximately 35% to human renin, Plasmepsin II was not inhibited significantly by renin inhibitors.
Hill
J
J
Department of Biochemistry, University of Wales College of Cardiff, UK.
Tyas
L
L
Phylip
L H
LH
Kay
J
J
Dunn
B M
BM
Berry
C
C
D000595:Amino Acid Sequence; D000818:Animals; D016282:Aspartic Acid Endopeptidases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D017125:Genes, Protozoan / Q000235:genetics; D007700:Kinetics; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000138:chemical synthesis / Q000378:metabolism / Q000494:pharmacology; D010963:Plasmodium falciparum / Q000201:enzymology* / Q000235:genetics; D011498:Protein Precursors / Q000378:metabolism; D015800:Protozoan Proteins; D011993:Recombinant Fusion Proteins / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000378:metabolism; D017421:Sequence Analysis; D013379:Substrate Specificity
D009842:Oligopeptides; D011498:Protein Precursors; D015800:Protozoan Proteins; D011993:Recombinant Fusion Proteins; D016282:Aspartic Acid Endopeptidases; C100637:plasmepsin II