Purification of Soluble and Membrane Forms of Somatic Angiotensin-Converting Enzyme by Cascade Affinity Chromatography

O. A. Kost,^1,2 S. V. Grinshtein,¹ I. I. Nikolskaya,¹ A. A. Shevchenko,¹ and P. V. Binevski¹

¹School of Chemistry, Lomonosov Moscow State University, Moscow, 119899 Russia; fax: (7-095) 939-35-89; E-mail: kost@enzyme.chem.msu.su

²To whom correspondence should be addressed.

Submitted November 10, 1996.

Soluble and membrane forms of angiotensin-converting enzyme were purified by cascade affinity chromatography. The enzyme forms were completely separated from each other using their different affinity to the hydrophobic matrix phenyl-silochrome. The enzymes were further purified by affinity sorbent prepared by immobilization of the enzyme inhibitor N-[1(S)-carboxy-5-aminopentyl]glycylphenylalanine. The procedure yielded electrophoretically homogeneous soluble and membrane forms of angiotensin-converting enzyme containing only active molecules as demonstrated by titration with the reversible inhibitor lisinopril. According to phase partition in the presence of Triton X-114, the membrane enzyme is more hydrophobic than the soluble form. The catalytic characteristics of the enzyme forms differed from each other in the system Aerosol OT--water--octane (reversed micelles) which is a model of the membrane environment of the enzymes in vivo.

KEY WORDS: angiotensin-converting enzyme, soluble form, membrane form, hydrophobic chromatography, affinity chromatography, Aerosol OT.

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