Reversible MM-creatine kinase binding to cardiac myofibrils

Abstract

Skinned rat papillary muscles and purified preparations of rat cardiac myofibrils were used to study the nature of the interaction of creatine kinase with cardiac myofibrils. High activity of creatine kinase (2 IU/mg protein in fibers and 0.9 IU/mg in purified myofibrils) was due mostly to reversibly bound enzyme. This activity could be removed and rebound. The process of creatine kinase rebinding was characterized by apparent Km value of 0.14 mg/ml (approximately equal to 2 X 10(6) M). Rebinding of creatine kinase to cardiac myofibrils restored the phenomenon of functional compartmentation of adenine nucleotides in myofibrillar space and restored the ability of phosphocreatine to decrease the rigor tension in the presence of MgADP. The physiological experiments with quick length changes showed that rebinding of creatine kinase to skinned papillary muscle also restored Ca sensitivity, increased maximal tension development, decreased stiffness, and restored the tension recovery after quick length changes in muscle under condition of inhibition of endogenous creatine kinase by 1-fluoro-2,4-dinitrobenzene. It is concluded that creatine kinase reversibly bound to cardiac myofibrils is involved in the energy supply for cardiac contraction.

An example of substrate channeling between co‐immobilized enzymes Coupled activity of myosin ATPase and creatine kinase bound to frog heart myofilaments

“…Under these conditions, 90% of the total CK activity was obtained in the supernates of the silicone oil centrifugation. It is very likely that some loosely bound CK [12] became detached from the myofilaments at this higher pH. In the bottom myofilament fraction, no CK activity was apparent by means of the ATP detection in the coupled enzymatic assay ( fig.2).…”

Total Creatine Kinase and Its Isoenzymes in Cord Blood of Full Term Newborns

“…The heart is characterized by having different CK isoenzymes (MBCK and mitochondrial (MtCK)) which are found specifically localized at sarcoplasmic reticulum, plasma membrane, myofilaments, mitochondria and glycolytic complexes (39,40).…”

The decrease of NAD(P)H:quinone oxidoreductase 1 activity and increase of ROS production by NADPH oxidases are early biomarkers in doxorubicin cardiotoxicity

“…The myofibril-containing pellet from the previous 1,000 x g centrifugation step was resuspended in myofibrils isolation buffer (Tris-HCl 20 mM pH 7.4, EDTA 1 mM and KCl 100 mM) containing 2% Triton X-100 (Ventura-Clapier et al 1987;Chicco et al 2006). Myofibrils were pellet down by centrifugation for 10 min at 6,000 x g and the treatment with Triton X-100 was repeated again.…”