A New Method for Measuring Serum Pyruvate Kinase and Creatine Kinase Activities Using a Thermostable Glucokinase

Abstract

A new method for measuring the activity of serum pyruvate kinase (PK: EC2. 7. 1. 40) was developed by using a thermostable glucokinase (Glck) from a thermophilic bacterium, Bacillus stearothermophilus. The ATP formed by the reaction of PK is finally converted to NADPH via glucose-6-phosphate by the action of Glck and glucose-6-phosphate dehydrogenase. The change in absorbance at 340nm was found to be linear up to about 5000U/I of PK. The within-run and day-to-day coefficients of variation were1.13% at 85.4U/l and 2.08% at 45.7U/l, respectively. The influence of various coexistents and anticoagulants, such as bilirubin, ascorbate, glucose, EDTA, sodium fluoride, etc., on the assay was negligible. The reagent was stable in solution for about one month at 10°.
A method for simultaneously measuring the activities of PK and creatine kinase (CK) in a single specimen was also developed. This was based on the fact that the assay conditions for both enzymes were similar. This method was found to have a high degree of precision and a good correlation with respective PK and CK assay methods. This simultaneous measurement may be useful for the accurate differential diagnosis of myocardial infarction.