Maldonado E, Soriano-García M, Moreno A, Cabrera N, Garza-Ramos G, de Gómez-Puyou M, Gómez-Puyou A, Perez-Montfort R
Departamento de Genética Molecular, Instituto de Química, UniversidadNacional Autónoma de México 4510 México.
J Mol Biol. 1998;283(1):193-203. doi: 10.1006/jmbi.1998.2094.
The aligned amino acid sequences of TIM from Trypanosoma cruzi (TcTIM) and Trypanosoma brucei (TbTIM) have a positional identity of 68%. The two enzymes have markedly similar catalytic properties. Agents that interact with their interface Cys inhibit TcTIM and TbTIM; and those TIMs that lack this Cys (such as human TIM) are largely or completely insensitive to these agents. The susceptibility of TcTIM to the agents is approximately 100 times higher than that of TbTIM. To ascertain the cause of this large difference, the crystal structure of TcTIM was solved at 1.83 A resolution. The two enzymes are very similar homodimers. In TcTIM and TbTIM their respective Cys, 15 or 14, forms part of the dimer interface. In both, the contacts of the Cys with residues of the other subunit are almost identical. Nevertheless, there are noteworthy differences between the two; the existence of glutamine 18 in TbTIM instead of glutamic acid in TcTIM at the beginning of helix 1 decreases the contacts between this portion of the protein and helix 3 of the other subunit. In addition, TcTIM has proline at position 24 in the first helix of the TIM barrel; this is absent in the other TIM. Pro24 disrupts the regular helix arrangement, making the pitch of this helix 1.2 A longer than in TbTIM. When Pro24 of TcTIM was substituted for Glu, the sensitivity of TcTIM to sulfhydryl reagents increased about fivefold, possibly as a consequence of an increase in the space between the first portion of helix 1 and helix 3 of the other subunit. Therefore, it may be concluded that the geometry of the latter region is central in the accessibility to agents that perturb the interface Cys. In human TIM this region is more compact.
克氏锥虫（TcTIM）和布氏锥虫（TbTIM）的TIM氨基酸序列比对后的位置一致性为68%。这两种酶具有明显相似的催化特性。与它们的界面半胱氨酸相互作用的试剂会抑制TcTIM和TbTIM；而那些缺乏这种半胱氨酸的TIM（如人类TIM）对这些试剂基本不敏感或完全不敏感。TcTIM对这些试剂的敏感性比TbTIM高约100倍。为了确定这种巨大差异的原因，以1.83埃的分辨率解析了TcTIM的晶体结构。这两种酶都是非常相似的同型二聚体。在TcTIM和TbTIM中，它们各自的半胱氨酸（15或14）构成了二聚体界面的一部分。在两者中，半胱氨酸与另一个亚基的残基的接触几乎相同。然而，两者之间存在值得注意的差异；在螺旋1起始处，TbTIM中的谷氨酰胺18取代了TcTIM中的谷氨酸，这减少了该蛋白质部分与另一个亚基的螺旋3之间的接触。此外，TcTIM在TIM桶第一个螺旋的第24位有脯氨酸；其他TIM中没有。Pro24破坏了规则的螺旋排列，使该螺旋的螺距比TbTIM中的长1.2埃。当将TcTIM的Pro24替换为Glu时，TcTIM对巯基试剂的敏感性增加了约五倍，这可能是由于螺旋1的第一部分与另一个亚基的螺旋3之间的空间增加所致。因此，可以得出结论，后一个区域的几何结构对于干扰界面半胱氨酸的试剂的可及性至关重要。在人类TIM中，这个区域更紧凑。
