Hodgkinson J L, Newman T M, Marston S B, Severs N J
Department of Cardiac Medicine, National Heart and Lung Institute, London, United Kingdom.
J Struct Biol. 1995 Mar-Apr;114(2):93-104. doi: 10.1006/jsbi.1995.1009.
The structure of the smooth muscle contractile apparatus was studied using ultrarapid freezing followed by freeze substitution or by longitudinal freeze-fracture, deep-etch, and platinum-carbon replication. Freeze substitution minimises the detrimental effects of chemical fixation and freeze fracture eliminates them entirely whilst revealing the ultrastructure in three dimensions. Unidirectionally shadowed freeze-fracture replicas of ultrarapidly frozen, relaxed, intact smooth muscle showed a well-preserved actin filament structure the 5.5-nm repeat of the actin subunits was clearly observed. In transversely fractured tissue the thick filaments were revealed, with a distribution comparable to that seen in transverse sections of freeze-substituted muscle. Relaxed muscle permeabilised using Triton X-100 showed a similar structure to that of intact tissue after ultrarapid freezing and examination both by freeze fracture and by freeze examination both by freeze fracture and by freeze substitution; the ratios of actin to myosin were also comparable. In permeabilised, rigorised tissue the structure of the actomyosin complex was revealed in detail; this was especially clear in freeze-substituted muscle. A cross-bridge spacing of 38 nm was measured in freeze-fractured, deep-etched tissue. The structural detail revealed is compatible with a side polar model of the actomyosin interaction and with the sliding filament mechanism of muscle contraction.
利用超快速冷冻后进行冷冻置换，或通过纵向冷冻断裂、深度蚀刻和铂 - 碳复型技术，对平滑肌收缩装置的结构进行了研究。冷冻置换可将化学固定的有害影响降至最低，而冷冻断裂则能完全消除这些影响，同时还能从三维角度揭示超微结构。对超快速冷冻、松弛、完整的平滑肌进行单向阴影冷冻断裂复型，显示出保存良好的肌动蛋白丝结构，清晰观察到肌动蛋白亚基的5.5纳米重复结构。在横向断裂的组织中，显示出粗丝，其分布与冷冻置换肌肉横切面中所见的分布相当。用Triton X - 100使松弛的肌肉透化后，经超快速冷冻并通过冷冻断裂和冷冻置换检查，其结构与完整组织相似；肌动蛋白与肌球蛋白的比例也相当。在透化、僵直的组织中，详细揭示了肌动球蛋白复合物的结构；这在冷冻置换的肌肉中尤为明显。在冷冻断裂、深度蚀刻的组织中测得横桥间距为38纳米。所揭示的结构细节与肌动球蛋白相互作用的侧极模型以及肌肉收缩的滑行丝机制相符。
