Differentiation by preparative continuous free flow-isoelectric focusing of cyclosporin A inhibitable peptidyl-prolyl cis/trans isomerase of human erythrocytes.
Preparative continuous free flow-isoelectric focusing has been used to separate at least three different components of intrinsic peptidyl-prolyl cis/trans isomerase (PPIase) activity from erythrocytes lysate. By adding chemical spacer molecules like glycine and Bicine to commercial carrier ampholyte mixtures the resulting pH profile was predictably influenced. With an applied field strength of 125-170 V/cm a residence time of less than 15 min was sufficient for the separation of PPIases with isoelectric points of 5.4, 5.7 and 5.9 from the bulky hemoglobin. The recovery of the overall PPIase activities was about 100%. The purification factor has been determined as 20- to 100-fold. For each isoform of the enzyme the peptidyl-prolyl cis/trans isomerase activity of the separated proteins was inhibited by cyclosporin A but was resistant toward FK 506.
Küllertz
G
G
Max-Planck-Gesellschaft, Arbeitsgruppe Enzymologie der Peptidbindung, Halle/Saale, Germany.
Meyer
S
S
Fischer
G
G
D000591:Amino Acid Isomerases / Q000037:antagonists & inhibitors / Q000302:isolation & purification*; D000665:Ampholyte Mixtures; D002352:Carrier Proteins / Q000037:antagonists & inhibitors / Q000302:isolation & purification*; D016572:Cyclosporine / Q000494:pharmacology*; D004912:Erythrocytes / Q000187:drug effects* / Q000201:enzymology; D006454:Hemoglobins / Q000302:isolation & purification; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007525:Isoelectric Focusing / Q000379:methods*; D019696:Peptidylprolyl Isomerase
D000665:Ampholyte Mixtures; D002352:Carrier Proteins; D006454:Hemoglobins; D016572:Cyclosporine; D000591:Amino Acid Isomerases; D019696:Peptidylprolyl Isomerase