[Affinity chromatography of proteases using peptide methyl ketones as ligands].
Peptide methyl ketones acting as reversible inhibitors of serine and cysteine proteases were introduced as a new type of ligands in the affinity chromatography. Subtilisins and related serine proteases with an inhibition constant to the soluble inhibitor Z-Ala2-PheCH3 of less or equal than 3 x 10(-5) M are nearly completely adsorbed by the affinity gel Sepharose 4B-epsilon-aminocaproic acid-Ala2-PheCH3. The well-known cysteine protease papain binds also to this gel. The elution of the enzymes could be performed by means of iso-propanol. Thermitase purified with this method is, regarding to its quality, comparable with enzyme preparations using isoelectric focusing and adsorption at porous glass bodies, respectively.
Institut für Biochemie, Bereich Medizin, Martin-Luther-Universität Halle-Wittenberg, Saale, DDR.
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Fittkau
D000433:1-Propanol; D000327:Adsorption; D000590:Amino Acid Chloromethyl Ketones / Q000378:metabolism*; D002846:Chromatography, Affinity / Q000379:methods*; D019286:Endopeptidase K; D010450:Endopeptidases / Q000302:isolation & purification; D005898:Glass; D007525:Isoelectric Focusing; D008024:Ligands; D010206:Papain / Q000302:isolation & purification; D010447:Peptide Hydrolases / Q000302:isolation & purification*; D011480:Protease Inhibitors / Q000378:metabolism*; D012697:Serine Endopeptidases / Q000302:isolation & purification; D012997:Solvents; D013381:Subtilisins / Q000302:isolation & purification
D000590:Amino Acid Chloromethyl Ketones; D008024:Ligands; D011480:Protease Inhibitors; D012997:Solvents; C047499:benzyloxycarbonylalanyl-alanyl phenylalanine chloromethyl ketone; D000433:1-Propanol; D010450:Endopeptidases; D010447:Peptide Hydrolases; D012697:Serine Endopeptidases; D013381:Subtilisins; C022728:thermitase; D019286:Endopeptidase K; D010206:Papain
Affinitätschromatographie von Proteasen mit Peptidmethylketonen als Liganden.