Modification of myofibrils by fluorophore-induced photo-oxidation.
The excitation of fluorophores in the vicinity of a myofibril stops both shortening in the presence of ATP and Ca2+, and the extraction of the A-band by NaCl in the presence of Mg pyrophosphate. Shortening is more quickly affected than extraction. These effects can be induced by fluorescently-tagged antibodies bound in the A-band. Both depolymerization of the thick filament and the interaction between the myosin head and actin appear to be modified. Enzymatic lowering of the oxygen concentration in the bathing solution during excitation reduces these effects, indicating that they are due to photo-oxidation catalysed by excitation of the fluorophore. The results suggest that care needs to be exercised to minimize the consequences of these changes on the outcome of fluorescence-based assays of activity. Irradiated myofibrils that do not shorten, hydrolyse ATP at a rate comparable to those that contract, so they may be useful as a model system for the study of crossbridge activity in the ordered array of proteins of the myofibril.
Knight
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P
Muscle Biology Department, Agricultural and Food Research Council Institute of Food Research, Bristol Laboratory, Langford, Avon, UK.
Parsons
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N
D000199:Actins / Q000528:radiation effects; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D017301:Ca(2+) Mg(2+)-ATPase / Q000378:metabolism; D002118:Calcium / Q000378:metabolism; D000252:Calcium-Transporting ATPases / Q000378:metabolism; D019793:Fluorescein; D005452:Fluoresceins / Q000494:pharmacology; D005455:Fluorescent Antibody Technique; D005456:Fluorescent Dyes / Q000494:pharmacology*; D008027:Light; D008856:Microscopy, Fluorescence / Q000379:methods*; D009119:Muscle Contraction / Q000187:drug effects* / Q000528:radiation effects; D009210:Myofibrils / Q000187:drug effects* / Q000528:radiation effects; D010084:Oxidation-Reduction; D010100:Oxygen / Q000378:metabolism; D010777:Photochemistry; D011817:Rabbits; D012235:Rhodamines / Q000494:pharmacology*; D014466:Ultraviolet Rays
D000199:Actins; D005452:Fluoresceins; D005456:Fluorescent Dyes; D012235:Rhodamines; D000255:Adenosine Triphosphate; D017301:Ca(2+) Mg(2+)-ATPase; D000252:Calcium-Transporting ATPases; D010100:Oxygen; D002118:Calcium; D019793:Fluorescein