A Protein That Binds Specifically to the M-Line of Skeletal Muscle Is Identified as the Muscle Form of Creatine Kinase

Abstract

Published information on the properties of two proteins from chicken muscle, creatine kinase (MMcreatine kinase) and an M-line protein, suggested that they might be identical molecules. Different published procedures were used to purify the two proteins to homogeneity, and the properties of the two preparations were compared. Creatine kinase specific activity increased during purification of M-line protein, reaching a value comparable to that of purified MM-creatine kinase. The two proteins migrated identically in two electrophoretic systems and, after electrophoresis, both could be stained for creatine kinase activity. Double immunodiffusion tests with antibody prepared against MM-creatine kinase established the serological identity of the two protein preparations. Immunofluorescent studies showed that antiserum against MM-creatine kinase was bound in a regular pattern at the centers of the A-band regions of isolated myofibrils. These data show conclusively that the M-line protein and MM-creatine kinase are identical.The major proteins of cross-striated skeletal muscle myofibrils (e.g., myosin, actin, tropomyosin) have been extensively investigated in recent years; their known interactions form the basis of our understanding of muscle contraction. Lately, attention has turned increasingly to the "minor" components of the contractile apparatus, those remaining proteins that may have structural, regulatory, and/or catalytic functions.One approach has been to focus on proteins that regulate activities associated with the contractile mechanism (e.g., Mg+2-stimulated adenosine triphosphatase) and to resolve, isolate, and characterize these factors (1, 2). Another approach has been to try to localize known proteins within myofibrils, by selective extraction, immunofluorescence, and histochemistry. As an example, there is evidence that some of the creatine kinase within muscle cells may be bound to myofibrils (3-7), though most of the creatine kinase appears to be soluble, located in the intermyofibrillar space (8, 9). A third approach has involved the localization, within the myofibrillar structure, of proteins for which no biological activity is yet known. Among the proteins studied in this way is one that evidently is localized at the M-line within the A-band region of each sarcomere (10, 11).Starting with a low ionic strength extraction of chicken skeletal muscle, Morimoto and Harrington (11) were able to purify this M-line protein to apparent homogeneity. These authors showed that the purified protein is a dimeric molecule of molecular weight 88,000, and they also determined its aminoacid composition.Comparison of these published data for M-line protein with the known data on molecular weight, subunit structure, extractability in low ionic strength, and aminoacid composition of the muscle (MM) form of creatine kinase (Table 1) suggested to us that these two muscle proteins might be identical. We have, therefore, purified M-line protein by the method of Morimoto and Harrington (11) and compared it with MM-...

Flight muscle function in Drosophila requires colocalization of glycolytic enzymes.

“…Our results showing that glycolytic enzymes are colocalized in a pattern with the primary site being Z-discs and M-lines is parallel to a set of studies on creatine kinase in vertebrates (Turner et al, 1973;Walliman et al, 1977a,b) and the invertebrate analogue of creatine kinase, arginine kinase, in Drosophila (Lang et al, 1980). These enzymes are also related to ATP metabolism (Van Waarde et al 1990).…”

Cytoarchitectural and metabolic adaptations in muscles with mitochondrial and cytosolic creatine kinase deficiencies

“…In muscle, MM-CK mediated ATP production is mainly coupled to the local activity of SR-and plasma-bound Ca 2+ -ATPases [6,8,9,32], the Na + /K + -ATPase [33,34] and the myosin ATPase involved in actin-myosin sliding during contraction in the sarcomeric M-band [35][36][37][38][39]. The MM-CK mediated reaction is also topologically coupled to the glycolytic metabolic reaction cascade in the I-band thinfilament region [35].…”

Isoform-specific Interaction of the Myosin-binding Proteins (MyBPs) with Skeletal and Cardiac Myosin Is a Property of the C-terminal Immunoglobulin Domain

“…These include titin or connectin (6,7), MyBP-C (C-protein) (8), MyBP-H (H-protein) (9,10), myomesin (11), M protein (12,13), skelemin (14), MM-creatine kinase (15,16), AMP-deaminase (17), and X-protein (the slow-type isoform of MyBP-C) (18 -20). The MyBPs, first isolated in crude myosin preparations by Offer and colleagues (21) in the early 1970s are a group of proteins distributed in the central two-thirds of the cross-bridge bearing region (C-zone) of the A-band (22).…”