A Specific Sequence of the Laminin 2 Chain Critical for the Initiation of Heterotrimer Assembly (*)
A Specific Sequence of the Laminin 2 Chain Critical for the Initiation of Heterotrimer Assembly (*)
复制标题
层粘连蛋白 2 链的特定序列对于异源三聚体组装的启动至关重要 (*)
DOI：
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发表时间：
1995
影响因子：
4.8
通讯作者：
Yoshihiko Yamada
中科院分区：
文献类型：
--
作者：
A. Utani;M. Nomizu;S. Sugiyama;S. Miyamoto;P. Roller;Yoshihiko Yamada
三链层粘连蛋白分子通过α-螺旋卷曲螺旋结构组装，每条链跨越约600个氨基酸残基。我们报道了β1和β 1链的C末端指导特异性二聚体和三聚体组装（Utani，A.，Nomizu，M.，廷普尔河，Roller，P. P.，Yamada，Y.（1994）J.Biol.Chem.269，19167-19175）。在这项研究中，我们专注于利用三种不同的方法的α2链的三聚体形成的机制。首先，使用突变的重组α2链的竞争测定将长臂C末端的25个氨基酸序列确定为与β1和β 1链组装的必需位点。该位点的定点突变和合成肽表明，该位点内的带正电荷的氨基酸残基和α-螺旋结构都是关键的。第二，重组α2链长臂的过表达研究证实，C-末端对于NIH 3 T3细胞内的三聚体组装是关键的。第三，圆二色光谱检测的复合物在体外重组揭示了动态构象变化的α2和1链的组装过程中。这些研究还表明，α2链末端C端的正确折叠对三聚体的稳定性至关重要。从这些数据可以得出结论，α2链长臂的C末端是有效启动层粘连蛋白异源三聚体组装所必需的。
Triple-stranded laminin molecules assemble via an α-helical coiled-coil structure spanning approximately 600 amino acid residues of each chain. We reported that the C termini of the β1 and 1 chains direct the specific dimer and trimer assembly (Utani, A., Nomizu, M., Timpl, R., Roller, P. P., and Yamada, Y.(1994) J. Biol. Chem. 269, 19167-19175). In this study, we focused on the mechanism of trimer formation of the α2 chain utilizing three different approaches. First, competition assays using mutated recombinant α2 chain defined a 25-amino acid sequence at the C terminus of the long arm as an essential site for assembly with β1 and 1 chain. Site-specific mutations and synthetic peptides of this site revealed that both positively charged amino acid residues and the α-helical structure within this site were critical. Second, overexpression studies of recombinant α2 chain long arm confirmed that the C-terminal end was critical for the trimer assembly within NIH 3T3 cells. Third, circular dichroism spectroscopic examination of the complexes reconstituted in vitro revealed dynamic conformational changes of the α2 and 1 chains in the process of assembly. These studies also revealed that the proper folding of the extreme C terminus of α2 chain was critical for the stability of trimer. From these data, it is concluded that the C terminus of α2 chain long arm is required for the effective initiation of laminin heterotrimer assembly.
DOI：
10.1073/pnas.87.9.3264
发表时间：
1990-05-01
影响因子：
11.1
作者：
EHRIG, K;LEIVO, I;ENGVALL, E
通讯作者：
ENGVALL, E
影响因子：
56.9
作者：
HARBURY, PB;ZHANG, T;ALBER, T
通讯作者：
ALBER, T