Efficient high-performance liquid chromatographic system for protein purification.
An efficient high-performance liquid chromatographic system, consisting of an affinity column and a high-performance size-exclusion column, was developed and applied to the purification of growth hormone receptors from rabbit livers. When a 6-ml sample of Triton X-100 extracts containing 16 mg of protein was applied to the system, 1200-fold purified receptor with a 10% recovery of binding activity from homogenates was obtained within 3-4 h. The purified receptor exhibited one main band on sodium dodecyl sulphate polyacrylamide gel electrophoresis, and the affinity constant (Ka = 6.0.10(9) M-1) was found to be comparable with that of 1% Triton X-100 extract (4.4.10(9) M-1). The injection of 1 ml of 3 M urea solution prior to receptor elution with 10 ml of 6 M urea solution was effective in removing non-specific binding proteins.
Yagi
S
S
Faculty of Pharmaceutical Sciences, Kyoto University, Japan.
Izawa
K
K
Nakagawa
T
T
Tanaka
H
H
Yoshitake
A
A
Mohri
Z
Z
D000818:Animals; D002846:Chromatography, Affinity; D002851:Chromatography, High Pressure Liquid / Q000295:instrumentation*; D004591:Electrophoresis, Polyacrylamide Gel; D008099:Liver / Q000378:metabolism; D011506:Proteins / Q000302:isolation & purification*; D011817:Rabbits; D011986:Receptors, Somatotropin / Q000302:isolation & purification; D013056:Spectrophotometry, Ultraviolet
D011506:Proteins; D011986:Receptors, Somatotropin