SUBUNIT STRUCTURE AND ASSEMBLY OF THE GLOBULAR DOMAIN OF BASEMENT-MEMBRANE COLLAGEN TYPE-IV
SUBUNIT STRUCTURE AND ASSEMBLY OF THE GLOBULAR DOMAIN OF BASEMENT-MEMBRANE COLLAGEN TYPE-IV
复制标题
DOI：
10.1111/j.1432-1033.1984.tb08019.x
复制
发表时间：
1984-01-01
期刊：
影响因子：
--
通讯作者：
TIMPL, R
中科院分区：
文献类型：
--
作者：
WEBER, S;ENGEL, J;TIMPL, R
用胶原酶溶出小鼠肿瘤、人胎盘和牛主动脉中的IV型胶原球状结构域，并用层析方法纯化。该材料显示出独特的非胶原氨基酸组成，并含有少量的氨基葡萄糖和半乳糖胺。分子量为17万的球状结构表现为由2个胶原IV分子组成的六聚体。该组合的亚基是两种不同的二聚体Da和Db （MW约56,000）和单体（MW = 28,000）。它们的n端氨基酸序列以短的三螺旋序列开始，与。α的c端三螺旋重叠。1（IV）和。alpha。2（IV）链，证明该小球起源于胶原IV的C端。二聚体是由单体通过二硫交联（形式Db）和/或不可还原交联（形式Da）形成的。在非变性条件下的还原会导致胶体和IV型胶原二聚体的部分解离，这表明两种不同IV型胶原分子的单体之间形成了可还原的交联。在8 M尿素、十二烷基硫酸钠或pH为2.5-4的条件下，六聚体可以解离成亚基。羧基显然是缔合所必需的。亚基（单体和二聚体）或纯化二聚体的混合物在中性缓冲液中重新组装成六聚体，如超离心和电子显微镜所示。然而，重组的六聚体在比天然球更宽的pH范围内解离。圆二向色光谱表明，酸处理的材料比尿素处理的材料结构更完全地折叠。源自单体的小球体（如存在于单一胶原IV分子中）显然被相邻的三螺旋结构稳定。共价交联的形成稳定了球状结构，并允许按化学计量比例重组。
The globular domain of collagen IV was solubilized by collagenase digestion from a mouse tumor, human placenta and bovine aorta and was purified by chromatographic methods. The materials show a unique, mainly non-collagenous amino acid composition and contain small amounts of glucosamine and galactosamine. The globular structures with MW = 170,000 appear as a hexameric assembly originating from 2 collagen IV molecules. Subunits of this assembly are 2 different dimers Da and Db (MW about 56,000) and monomers (MW = 28,000). Their N-terminal amino acid sequences start with short triple-helical sequences, which overlap with the C-terminal triple helix of the .alpha.1(IV) and .alpha.2(IV) chain, demonstrating that the globule originates from the C terminus of collagen IV. Dimers arise from monomers by disulfide cross-linking (form Db) and/or formation of non-reducible cross-links (form Da). Reduction under non-denaturing conditions causes partial dissociation of the globule and of collagen IV dimers, indicating that reducible cross-links are formed between monomers of 2 different collagen IV molecules. Dissociation of the hexamer into the subunits can be achieved with 8 M urea, sodium dodecyl sulfate or in the pH range 2.5-4. Carboxyl groups apparently are essential for association. Mixtures of the subunits (monomers and dimers) or purified dimers reassembled in neutral buffer into hexamers as shown by ultracentrifugation and EM. Reconstituted hexamers, however, dissociate in a much broader pH range than the native globules. Circular dichroic spectra indicate that the structure is more completely refolded from acid-treated than from urea-treated material. Globules originating from monomers (as existing in single collagen IV molecules) apparently are stabilized by the adjacent triple helix. Covalent cross-link formation stabilizes the globular structure and allows reconstitution in stoichiometric proportions.