The purification of human urinary kallikrein with ion-exchange radial flow membrane chromatography.
A method was developed for the purification of kallikrein from human urine. The procedure consisted of three steps: ultradialysis, diethyl-(2-hydroxypropyl) aminoethyl (QAE) ion exchange radial flow membrane chromatography and affinity chromatography on aprotinin agarose. It is simple and suitable for large-scale purification. The purified product was checked by SDS-PAGE and matrix-assisted laser desorption:ionization mass spectrometery (MALDI). A single band with apparent molecular weight (MW) 42,000 in SDS-PAGE and a single monomer peak with MW33,000 in MALDI were observed, respectively. The biological activity tested by ELISA showed positive immunological identity of the purified product compared with human urinary kallikrein standard.
Wang
H
H
National Chromatographic R. and A. Center, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Peoples Republic of China.
Li
T
T
Zou
H
H
Zhang
Y
Y
Chao
J
J
Chao
L
L
D000595:Amino Acid Sequence; D002847:Chromatography, Agarose; D002852:Chromatography, Ion Exchange / Q000191:economics / Q000295:instrumentation*; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D005456:Fluorescent Dyes / Q000737:chemistry; D006801:Humans; D007610:Kallikreins / Q000302:isolation & purification* / Q000652:urine*; D008297:Male; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000737:chemistry; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014462:Ultrafiltration
D005456:Fluorescent Dyes; D009842:Oligopeptides; C025273:prolyl-phenylalanyl-arginine-4-methylcoumaryl-7-amide; D007610:Kallikreins