Elastic properties of titin filaments demonstrated using a "freeze-break" technique.
A "freeze-break" technique (Trombitás, K.: Acta Biochim. Biophys. Hung. 6:419-427, 1971) and immunoelectron microscopy were used to study the elastic properties of titin filaments. Small bundles of freshly prepared rabbit psoas muscle fibers were quickly frozen and broken under liquid nitrogen to fracture sarcomeres in planes perpendicular to the filament axis, in each of various regions along the sarcomere. The still-frozen specimens were thawed during fixation to allow elastic filaments to retract. The broken specimens were then labelled with monoclonal anti-titin antibodies against an unique epitope in the I-band. The titin epitopes were normally positioned symmetrically about the Z-line. However, in sarcomeres broken at the A-I junction, the epitopes no longer remained symmetrical: the titin filaments in the broken half-sarcomere retracted, independently of the thin filaments, forming a dense band just near the Z-line. The retracted density apparently did not reach the Z-line; retraction stopped at the level of the so-called N1-line. In sarcomeres broken at the Z-line level, the titin filaments retracted in the opposite direction. In this case the titin epitope retracted all the way to the ends of the thick filaments. It appears then that titin molecules form elastic filaments that are independent of thin filaments in most of the I-band. Near the Z-line, however, the titin filaments either have an inelastic domain or associate firmly with the thin filaments at the N1-line level.
Trombitás
K
K
Center for Bioengineering, University of Washington, Seattle.
Pollack
G H
GH
Wright
J
J
Wang
K
K
D008841:Actin Cytoskeleton / Q000648:ultrastructure*; D000818:Animals; D064211:Connectin; D004548:Elasticity; D005614:Freeze Fracturing; D016253:Microscopy, Immunoelectron; D009124:Muscle Proteins / Q000737:chemistry / Q000648:ultrastructure*; D011494:Protein Kinases; D011817:Rabbits; D012518:Sarcomeres / Q000648:ultrastructure
D064211:Connectin; D009124:Muscle Proteins; D011494:Protein Kinases