Production of recombinant human creatine kinase (r-hCK) isozymes by tandemrepeat expression of M and B genes and characterization of r-hCK-MB

Background: Serum creatine kinase-MB isoenzyme (CK-MB) is widely used as a marker of myocardial injury. We prepared recombinant human CK (r-hCK) MB is oenzyme and examined its potential for use as a control material for assay of CK-MB in serum. Methods: cDNAs encoding CK-M and CK-B subunits were inserted into the same plasmid vector, followed by transformation of Escherichia coli. The resulti ng three types of CK isoenzymes were purified by conventional chromatograph y. Results: The ratio of MB to MM to BE was 50:40:10 on the basis of CK activi ty. Highly purified CK-MB with a specific activity of 533 U/mg was produced in a yield of 5.7 mg/g of packed cells. Purified r-hCK-MB had the isoelect ric point (pI 5.3) and molecular size (46 kDa for the subunit) of native CK -MB. Its immunoreactivity in an ELISA using antibody against native heart e nzyme was similar to that of cardiac CK-MB. The r-hCK-MB retained >90% acti vity for at least 4 months at 11 degreesC in a delipidated serum matrix in a liquid form at a concentration of 118 U/L. Conclusions: r-hCK-MB shows key properties of the native cardiac isoenzyme and may be useful as a control and calibrator for serum assays of CK-MB. (C ) 2001 American Association for Clinical Chemistry.