Biochemical and ultrastructural aspects of Mr 165,000 M-protein in cross-striated chicken muscle.
To better understand the relationship between the Mr 165,000 M-line protein (M-protein) and H-zone structure in skeletal and in cardiac muscle, as well as the possible interaction of M-protein with another skeletal muscle M-line component, the homodimeric creatine kinase isoenzyme composed of two M subunits (MM-CK), we performed biochemical, immunological, and ultrastructural studies on myofibrils extracted by different procedures. In contrast to MM-CK, M-protein could not be completely removed from myofibrils by low ionic strength extraction. Fab-fragments of antibodies against M-protein could not release M-protein quantitatively from either breast or heart myofibrils but remained bound to the myofibrillar structure, whereas monovalent antibodies against MM-CK cause the specific release of MM-CK and the concomitant disappearance of the M-line from chicken skeletal muscle myofibrils. When MM-CK was removed from skeletal myofibrils by low ionic strength extraction or, more specifically, by incubation with anti-MM-CK Fab, M-protein was still not released quantitatively upon treatment with anti-M-protein Fab as judged from immunofluorescence data. In the ultrastructural investigation of low ionic strength extracted muscle fibers, M protein could be localized in two stripes on both sides of the former M-line, suggesting a reduced attachment to the residual H-zone structure, whereas the specific removal of MM-CK resulted in the same dense staining pattern for M-protein within the M-line as observed in untreated fibers. However, the binding of M-protein to the residual M-line structure seemed to be reduced, as a considerable amount of this protein could be identified in the supernate of sequentially incubated myofibrils. The results indicate a strong binding of M-protein within the H-zone structure of skeletal as well as heart myofibrils.
D000818:Animals; D002458:Cell Fractionation / Q000379:methods; D002645:Chickens; D003402:Creatine Kinase / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D007158:Immunologic Techniques; D008854:Microscopy, Electron; D008970:Molecular Weight; D009124:Muscle Proteins / Q000302:isolation & purification*; D009132:Muscles / Q000201:enzymology / Q000648:ultrastructure*; D009206:Myocardium / Q000648:ultrastructure*
D009124:Muscle Proteins; D003402:Creatine Kinase