Steinert P M, Steven A C, Roop D R
J Invest Dermatol. 1983 Jul;81(1 Suppl):86s-90s. doi: 10.1111/1523-1747.ep12540757.
We have studied the structure of epidermal keratin filaments polymerized in vitro, addressing two different levels of organization. First, we have determined the amino acid sequence of a mouse epidermal keratin subunit from the nucleotide sequence of a cDNA clone. The subunit contains a large central region, representing about 50 percent, whose sequence strongly suggests that it assumes a coiled-coil alpha-helical conformation. This is flanked on the amino and carboxyl terminals by long glycine-rich sequences. Second, we have used scanning transmission electron microscopy to study the structure of frozen, unstained filaments. Analyses of such images provides information on the mass per unit length and on the distribution of mass within the filament. These data impose rigorous constraints on possible models for the packing of protofilaments within the filament. Epidermal keratin filaments assembled in vitro are polymorphic; however, the majority of bovine filaments weigh about 37 kD/nm, but most human filaments have masses of only about 27 kD/nm. The filament width is at least 15 nm, substantially more than the generally accepted value of 8 to 10 nm, owing to the existence of low-density mass at the periphery that has not been visualized by conventional microscopic methods. We currently postulate that the alpha-helical regions of the subunits comprise the structural core or backbone of the filament from which at least some of the glycine-rich sequences protrude.
我们研究了在体外聚合的表皮角蛋白丝的结构，涉及两个不同的组织层次。首先，我们从一个cDNA克隆的核苷酸序列中确定了小鼠表皮角蛋白亚基的氨基酸序列。该亚基包含一个大约占50%的大的中央区域，其序列强烈表明它呈现出卷曲螺旋α-螺旋构象。在氨基和羧基末端两侧是富含甘氨酸的长序列。其次，我们使用扫描透射电子显微镜研究了冷冻、未染色的细丝的结构。对这些图像的分析提供了关于每单位长度的质量以及细丝内质量分布的信息。这些数据对细丝内原纤维堆积的可能模型施加了严格的限制。体外组装的表皮角蛋白丝是多态的；然而，大多数牛细丝的重量约为37 kD/nm，但大多数人细丝的质量仅约为27 kD/nm。细丝宽度至少为15 nm，大大超过了通常接受的8至10 nm的值，这是由于周边存在低密度物质，而传统显微镜方法无法观察到这些物质。我们目前推测，亚基的α-螺旋区域构成了细丝的结构核心或主干，至少一些富含甘氨酸的序列从该核心或主干突出。
