Substrate channelling in a XXXX kinase system of rat skeletal muscle under various pH conditions.

@entity908 @entity857 @entity855
The aim of this study was to evaluate myofibrillar @entity908 kinase (CK) activity and to quantify the substrate channelling of @entity855 between CK and myosin ATPase under different pH conditions within the integrity of myofibrils. A pure myofibrillar fraction was prepared using differential centrifugation. The homogeneity of the preparation and the purity of the fraction were confirmed microscopically and by enzymatic assays for contaminant enzyme activities. The specific activity of myofibrillar CK reached 584 +/- 33 nmol PCr min(-1) mg(-1) at pH 6.75. Two methods were used to detect CK activity: (1) measurement of direct @entity855 production, and (2) measurement of PCr consumption. This method of evaluation has been tested in experiments with isolated @entity908 kinase. No discrepancy in CK activity between the methods was observed in the pH range tested (6.0-7.5). However, the same procedures resulted in a significant discrepancy between the amounts of reacted PCr and produced @entity855 within the pure myofibrillar fraction. This discrepancy represents the portion of @entity855 produced by the CK reaction, which is preferentially channelled to the myosin ATPase before diffusing into the bulk solution. The maximum evaluated difference reached 42.3 % at pH 6.95. The substrate channelling between myofibrillar-bound CK and myosin ATPase was evaluated under various pH levels within the physiological range and it reached a maximum value in a slightly acidic environment. These results suggest that @entity855 / @entity857 flux control by the CK system is more important at lower pH, corresponding to the physiological state of muscle fatigue.
