Purification and characterization of recombinant human cathepsin E.
The human cathepsin E was purified from the culture supernatant of Pichia pastoris strain transformed with a human cathepsin E expression plasmid. Purification was performed by a three-step procedure, TSKgel Phenyl-5PW, Toyopearl HW55S and TSKgel DEAE-5PW column chromatographies. The purified recombinant cathepsin E had the molecular mass of around 82-kDa with the amino-terminal sequence started with Ile37 of the predicted amino acid sequence, suggesting the human cathepsin E was accumulated in the culture suparnatant as the mature dimer enzyme. The result of endoglycosidase-H digestion followed by Western blot analysis of the purified recombinant cathepsin E suggested that the human cathepsin E expressed in Pichia pastoris received N-linked high-mannose type glycosylation.
Iida
H
H
Biotechnology Research Laboratory, Tosoh Corporation, Kanagawa, Japan.
Matsuba
T
T
Yamada
M
M
Azuma
T
T
Suzuki
H
H
Yamamoto
K
K
Hori
H
H
D020059:Cathepsin E; D002403:Cathepsins / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D003001:Cloning, Molecular; D003470:Culture Media; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008970:Molecular Weight; D010843:Pichia; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*
D003470:Culture Media; D011994:Recombinant Proteins; D002403:Cathepsins; D020059:Cathepsin E