X-RAY-DIFFRACTION INDICATES THAT ACTIVE CROSS-BRIDGES BIND TO ACTIN TARGET ZONES IN INSECT FLIGHT-MUSCLE

We report the first time-resolved study of the two-dimensional x-ray d iffraction pattern during active contraction in insect flight muscle ( IFM), Activation of demembranated Lethocerus IFM was triggered by 1.5- 2.5% step stretches (risetime 10 ms; held for 1.5 s) giving delayed ac tive tension that peaked at 100-200 ms. Bundles of 8-12 fibers were st retch-activated on SRS synchrotron x-ray beamline 16.1, and time-resol ved changes in diffraction were monitored with a SRS 2-D multiwire det ector. As active tension rose, the 14.5- and 7.2-nm meridionals fell, the first row line dropped at the 38.7 nm layer line while gaining a n ew peak at 19.3 nm, and three outer peaks on the 38.7-nm layer line ro se. The first row line changes suggest restricted binding of active my osin heads to the helically preferred region in each actin target zone , where, in rigor, two-headed lead bridges bind, midway between tropon in bulges that repeat every 38.7 nm, Halving this troponin repeat by b inding of single active heads explains the intensity rise at 19.3 nm b eing coupled to a loss at 38.7 nm, The meridional changes signal movem ent of at least 30% of all myosin heads away from their axially ordere d positions on the myosin helix. The 38.7- and 19.3-nm layer line chan ges signal stereoselective attachment of 7-23% of the myosin heads to the actin helix, although with too little ordering at 6-nm resolution to affect the 5.9-nm actin layer line. We conclude that stretch-activa ted tension of IFM is produced by cross-bridges that bind to rigor's l ead-bridge target zones, comprising less than or equal to 1/3 of the 7 5-80% that attach in rigor.