PMID:10320340

Citation	Luo, G, Herrera, AH and Horowits, R (1999) Molecular interactions of N-RAP, a nebulin-related protein of striated muscle myotendon junctions and intercalated disks. Biochemistry 38:6135-43
Abstract	N-RAP is a recently discovered muscle-specific protein that is concentrated at the myotendon junctions in skeletal muscle and at the intercalated disks in cardiac muscle. The C-terminal half of N-RAP contains a region with sequence homology to nebulin, while a LIM domain is found at its N-terminus. N-RAP is hypothesized to perform an anchoring function, linking the terminal actin filaments of myofibrils to protein complexes located beneath the sarcolemma. We used a solid-phase assay to screen myofibrillar and junctional proteins for binding to several recombinant fragments of N-RAP, including the nebulin-like super repeat region (N-RAP-SR), the N-terminal half including the LIM domain (N-RAP-NH), and the region of N-RAP between the super repeat region and the LIM domain (N-RAP-IB). Actin is the only myofibrillar protein tested that exhibits specific binding to N-RAP, with high-affinity binding to N-RAP super repeats, and 10-fold weaker binding to N-RAP-IB. In contrast, myosin, isolated myosin heads, tropomyosin, and troponin exhibited no specific interaction with N-RAP domains. A recombinant fragment corresponding to the C-terminal one-fourth of vinculin also binds specifically to N-RAP super repeats, while no specific N-RAP binding activity was observed for other regions of the vinculin molecule. Finally, talin binds with high affinity to the LIM domain of N-RAP. These results support our hypothesis that N-RAP is part of a complex of proteins that anchors the terminal actin filaments of the myofibril to the membrane, and functions in transmitting tension from the myofibrils to the extracellular matrix.
Links	PubMed Online version:10.1021/bi982395t
Keywords	Actins/metabolism; Animals; Antibodies; Cell Membrane/metabolism; Mice; Muscle Proteins/chemistry; Muscle Proteins/immunology; Muscle Proteins/metabolism; Muscles/metabolism; Myofibrils/metabolism; Osmolar Concentration; Sarcolemma/metabolism; Sarcolemma/physiology; Talin/metabolism; Vinculin/metabolism
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