Nakamura A, Arai T, Kondo Y, Fujii T
Department of Functional Polymer Science, Faculty of Textile Science and Technology, Shinshu University, Nagano.
J Biochem. 1989 Jul;106(1):93-7. doi: 10.1093/oxfordjournals.jbchem.a122827.
The effects of poly(L-aspartic acid) (PLAA) on microtubule assembly and microtubule-associated protein (MAP) 2-actin interaction were examined in vitro. PLAA inhibited assembly of rat brain microtubules and induced rapid disassembly of already formed microtubules. Inhibition was stronger by PLAA with a high molecular weight than that by low molecular weight. The ratios of 47 kDa PLAA to microtubule proteins causing 50% inhibition of the assembly and disassembly were 0.015 and 0.04 (w/w), respectively. Both MAP 1 and MAP 2 were bound to a PLAA-Sepharose 4B affinity column, while tubulin was not retained by the column. PLAA caused selective dissociation of MAP 1 and MAP 2 from microtubules polymerized by taxol. It is therefore concluded that PLAA interacts specifically with MAPs. PLAA also inhibited the MAP 2 induced cross-linking of actin filaments, showing an almost complete inhibition at a PLAA to MAP 2 ratio of 1:5,000 (w/w). Binding experiments of PLAA with digested MAP 2 by chymotrypsin using affinity chromatography and sedimentation experiments showed that PLAA was preferentially bound to a 35 kDa fragment which includes the microtubule- and actin-binding domain of the MAP 2 molecule. These results suggest that PLAA suppressed the functions of MAP 2 through a domain which is located in the 35 kDa fragment.
在体外研究了聚（L-天冬氨酸）（PLAA）对微管组装以及微管相关蛋白（MAP）2-肌动蛋白相互作用的影响。PLAA抑制大鼠脑微管的组装，并诱导已形成微管的快速解聚。高分子量的PLAA比低分子量的PLAA抑制作用更强。导致组装和解聚50%抑制的47 kDa PLAA与微管蛋白的比例分别为0.015和0.04（w/w）。MAP 1和MAP 2都与PLAA-琼脂糖4B亲和柱结合，而微管蛋白未被该柱保留。PLAA导致MAP 1和MAP 2从紫杉醇聚合的微管上选择性解离。因此得出结论，PLAA与微管相关蛋白特异性相互作用。PLAA还抑制了MAP 2诱导的肌动蛋白丝交联，在PLAA与MAP 2比例为1:5000（w/w）时几乎完全抑制。使用亲和色谱法和沉降实验对PLAA与胰凝乳蛋白酶消化的MAP 2进行结合实验表明，PLAA优先与一个包含MAP 2分子微管和肌动蛋白结合结构域的35 kDa片段结合。这些结果表明，PLAA通过位于35 kDa片段中的一个结构域抑制了MAP 2的功能。
