Johnson Kenneth A, Angelucci Francesco, Bellelli Andrea, Hervé Maxime, Fontaine Josette, Tsernoglou Demetrious, Capron André, Trottein François, Brunori Maurizio
Department of Biochemical Sciences and Istituto Pasteur-Fondazione Cenci Bolognetti, University of Rome La Sapienza, Rome, Italy,
Biochemistry. 2003 Sep 2;42(34):10084-94. doi: 10.1021/bi034449r.
Schistomiasis is a debilitating parasitic disease which affects 200 million people, causing life-threatening complications in 10% of the patients. This paper reports the crystal structure of the Schistosoma haematobium 28 kDa glutathione S-transferase, a multifunctional enzyme involved in host-parasite interactions and presently considered as a promising vaccine candidate against schistosomiasis. The structures of the GSH-free enzyme, as well as the partially (approximately 40%) and almost fully (approximately 80%) GSH-saturated enzyme, exhibit a unique feature, absent in previous GST structures, concerning the crucial and invariant Tyr10 side chain which occupies two alternative positions. The canonical conformer, which allows an H-bond to be formed between the side chain hydroxyl group and the activated thiolate of GSH, is somewhat less than 50% occupied. The new conformer, with the phenoxyl ring on the opposite side of the mobile loop connecting strand 1 and helix 1, is stabilized by a polar interaction with the guanidinium group of the conserved Arg21 side chain. The presence of two conformers of Tyr10 may provide a clue about clarifying the multiple catalytic functions of Sh28GST and might prove to be relevant for the design of specific antischistosomal drugs. The K(d) for GSH binding was determined by equilibrium fluorescence titrations to be approximately 3 microM and by stopped-flow rapid mixing experiments to be approximately 9 microM. The relatively tight binding of GSH by Sh28GST explains the residually bound GSH in the crystal and supports a possible role of GSH as a tightly bound cofactor involved in the catalytic mechanism for prostaglandin D(2) synthase activity.
血吸虫病是一种使人虚弱的寄生虫病，影响着2亿人，10%的患者会出现危及生命的并发症。本文报道了埃及血吸虫28 kDa谷胱甘肽S-转移酶的晶体结构，该酶是一种多功能酶，参与宿主与寄生虫的相互作用，目前被认为是一种有前景的抗血吸虫病疫苗候选物。无谷胱甘肽（GSH）的酶以及部分（约40%）和几乎完全（约80%）GSH饱和的酶的结构，展现出一个在先前谷胱甘肽S-转移酶（GST）结构中不存在的独特特征，即关键且不变的Tyr10侧链占据两个交替位置。允许侧链羟基与GSH的活化硫醇盐之间形成氢键的经典构象体占有率略低于50%。新的构象体中，苯氧基环位于连接链1和螺旋1的活动环的另一侧，通过与保守的Arg21侧链的胍基形成极性相互作用而得以稳定。Tyr10的两种构象体的存在可能为阐明Sh28GST的多种催化功能提供线索，并且可能证明与设计特定的抗血吸虫药物相关。通过平衡荧光滴定法测定GSH结合的解离常数（K(d)）约为3 microM，通过停流快速混合实验测定约为9 microM。Sh28GST对GSH的相对紧密结合解释了晶体中残留结合的GSH，并支持GSH作为参与前列腺素D2合酶活性催化机制的紧密结合辅因子的可能作用。
