Wierenga R K, Noble M E, Postma J P, Groendijk H, Kalk K H, Hol W G, Opperdoes F R
European Molecular Biology Laboratory, Heidelberg, Federal Republic of Germany.
Proteins. 1991;10(1):33-49. doi: 10.1002/prot.340100105.
Triosephosphate isomerase has an important loop near the active site which can exist in a "closed" and in an "open" conformation. Here we describe the structural properties of this "flexible" loop observed in two different structures of trypanosomal triosephosphate isomerase. Trypanosomal triosephosphate isomerase, crystallized in the presence of 2.4 M ammonium sulfate, packs as an asymmetric dimer of 54,000 Da in the crystallographic asymmetric unit. Due to different crystal contacts, peptide 167-180 (the flexible loop of subunit-1) is an open conformation, whereas in subunit-2, this peptide (residues 467-480) is in a closed conformation. In the closed conformation, a hydrogen bond exists between the tip of the loop and a well-defined sulfate ion which is bound to the active site of subunit-2. Such an active site sulfate is not present in subunit-1 due to crystal contacts. When the native (2.4 M ammonium sulfate) crystals are transferred to a sulfate-free mother liquor, the flexible loop of subunit-2 adopts the open conformation. From a closed starting model, this open conformation was discovered through molecular dynamics refinement without manual intervention, despite involving C alpha shifts of up to 7 A. The tip of the loop, residues 472, 473, 474, and 475, moves as a rigid body. Our analysis shows that in this crystal form the flexible loop of subunit-2 faces a solvent channel. Therefore the open and the closed conformations of this flexible loop are virtually unaffected by crystal contacts. The actual observed conformation depends only on the absence or presence of a suitable ligand in the active site.
磷酸丙糖异构酶在活性位点附近有一个重要的环，它可以以“闭合”和“开放”两种构象存在。在此，我们描述了在锥虫磷酸丙糖异构酶的两种不同结构中观察到的这个“柔性”环的结构特性。锥虫磷酸丙糖异构酶在2.4 M硫酸铵存在下结晶，在晶体学不对称单元中以54,000 Da的不对称二聚体形式堆积。由于不同的晶体接触，肽段167 - 180（亚基1的柔性环）处于开放构象，而在亚基2中，该肽段（残基467 - 480）处于闭合构象。在闭合构象中，环的末端与一个明确的硫酸根离子之间存在氢键，该硫酸根离子与亚基2的活性位点结合。由于晶体接触，亚基1中不存在这样的活性位点硫酸根。当天然（2.4 M硫酸铵）晶体转移到无硫酸根的母液中时，亚基2的柔性环会采用开放构象。从一个闭合的起始模型出发，尽管涉及高达7 Å的Cα位移，但通过分子动力学优化在没有人工干预的情况下发现了这种开放构象。环的末端，即残基472、473、474和475，作为一个刚体移动。我们的分析表明，在这种晶体形式中，亚基2的柔性环面向一个溶剂通道。因此，这个柔性环的开放和闭合构象实际上不受晶体接触的影响。实际观察到的构象仅取决于活性位点中是否存在合适的配体。
