Trask T M, Ritty T M, Broekelmann T, Tisdale C, Mecham R P
Department of Cell Biology and Physiology, Washington University School of Medicine, Box 8228, 660 South Euclid Avenue, St. Louis, MO 63110, USA.
Biochem J. 1999 Jun 15;340 ( Pt 3)(Pt 3):693-701.
Aggregation of fibrillin molecules via disulphide bonds is postulated to be an early step in microfibril assembly. By expressing fragments of fibrillin 1 and fibrillin 2 in a mammalian expression system, we found that the N-terminal region of each protein directs the formation of homodimers and that disulphide bonds stabilize this interaction. A large fragment of fibrillin 1 containing much of the region downstream from the N-terminus remained as a monomer when expressed in the same cell system, indicating that this region of the protein lacks dimerization domains. This finding also confirms that the overexpression of fibrillin fragments does not in itself lead to spurious dimer formation. Pulse-chase analysis demonstrated that dimer formation occurred intracellularly, suggesting that the process of fibrillin aggregation is initiated early after biosynthesis of the molecules. These findings also implicate the N-terminal region of fibrillin 1 and fibrillin 2 in directing the formation of a dimer intermediate that aggregates to form the functional microfibril.
据推测,原纤维蛋白分子通过二硫键聚集是微原纤维组装的早期步骤。通过在哺乳动物表达系统中表达原纤维蛋白1和原纤维蛋白2的片段,我们发现每种蛋白质的N端区域指导同型二聚体的形成,并且二硫键稳定了这种相互作用。当在相同的细胞系统中表达时,包含N端下游大部分区域的原纤维蛋白1的大片段仍为单体,这表明该蛋白质的这一区域缺乏二聚化结构域。这一发现还证实,原纤维蛋白片段的过表达本身不会导致假二聚体的形成。脉冲追踪分析表明二聚体形成发生在细胞内,这表明原纤维蛋白聚集过程在分子生物合成后早期就开始了。这些发现还表明原纤维蛋白1和原纤维蛋白2的N端区域在指导形成聚集形成功能性微原纤维的二聚体中间体方面发挥作用。