Hugdahl J D, Bokros C L, Hanesworth V R, Aalund G R, Morejohn L C
Department of Botany, University of Texas at Austin 78713.
Plant Cell. 1993 Sep;5(9):1063-80. doi: 10.1105/tpc.5.9.1063.
An understanding of the regulation of microtubule polymerization and dynamics in plant cells requires biochemical information on the structures, functions, and molecular interactions of plant tubulin and microtubule-associated proteins (MAPs) that regulate microtubule function. We have probed the regulatory domain and polymerization domain of purified maize tubulin using MAP2, an extensively characterized mammalian neuronal MAP. MAP2 bound to the surface of preformed, taxol-stabilized maize microtubules, with binding saturation occurring with one MAP2 molecule per five to six tubulin dimers, as it does with mammalian microtubules. MAP2 binding and dissociation analyses revealed two affinity classes of binding sites on maize microtubules: a high-affinity site 12 dimers apart that may be homologous to the mammalian MAP2 binding site and an additional low-affinity site also 12 dimers apart that may be homologous to the mammalian tau binding site. MAP2 corrected in vitro folding errors in taxol-stabilized maize microtubules and reduced the critical concentration of maize tubulin polymerization eightfold, from 8.3 to 1.0 microM. However, MAP2 dissociated much more readily from maize microtubules than from mammalian microtubules and induced the assembly of maize tubulin into aberrant helical ribbon polymers that remained stable for prolonged periods. Our results indicated that MAP2 binds to maize tubulin via a partially specific, low-fidelity interaction that reflects unique structural and functional properties of the polymerization and regulatory domains of plant tubulin and possibly of the tubulin binding domains of undocumented MAPs that regulate microtubule function in plant cells.
要了解植物细胞中微管聚合和动力学的调控，需要有关植物微管蛋白和微管相关蛋白（MAPs）的结构、功能及分子相互作用的生化信息，这些蛋白调控着微管功能。我们使用MAP2（一种已被广泛研究的哺乳动物神经元MAP）探究了纯化的玉米微管蛋白的调控结构域和聚合结构域。MAP2结合到预先形成的、紫杉醇稳定的玉米微管表面，结合饱和度为每五到六个微管蛋白二聚体有一个MAP2分子，与哺乳动物微管的情况相同。MAP2结合和解离分析揭示了玉米微管上存在两类亲和力不同的结合位点：一类高亲和力位点相隔12个二聚体，可能与哺乳动物MAP2结合位点同源；另一类低亲和力位点也相隔12个二聚体，可能与哺乳动物tau结合位点同源。MAP2纠正了紫杉醇稳定的玉米微管中的体外折叠错误，并将玉米微管蛋白聚合的临界浓度降低了八倍，从8.3微摩尔降至1.0微摩尔。然而，MAP2从玉米微管上解离比从哺乳动物微管上更容易，并且诱导玉米微管蛋白组装成异常的螺旋带状聚合物，这些聚合物能长时间保持稳定。我们的结果表明，MAP2通过部分特异性、低保真度的相互作用与玉米微管蛋白结合，这反映了植物微管蛋白聚合和调控结构域以及可能调控植物细胞微管功能的未记录MAPs的微管蛋白结合结构域的独特结构和功能特性。
