Wintjens René, Belrhali Hassan, Clantin Bernard, Azarkan Mohamed, Bompard Coralie, Baeyens-Volant Danielle, Looze Yvan, Villeret Vincent
Laboratoire de Chimie Générale, Institut de Pharmacie-U.L.B. CP 206/04, Boulevard du Triomphe, B-1050 Brussels, Belgium.
J Mol Biol. 2006 Mar 24;357(2):457-70. doi: 10.1016/j.jmb.2005.12.029. Epub 2006 Jan 4.
Glutaminyl cyclases (QCs) (EC 2.3.2.5) catalyze the intramolecular cyclization of protein N-terminal glutamine residues into pyroglutamic acid with the concomitant liberation of ammonia. QCs may be classified in two groups containing, respectively, the mammalian enzymes, and the enzymes from plants, bacteria, and parasites. The crystal structure of the QC from the latex of Carica papaya (PQC) has been determined at 1.7A resolution. The structure was solved by the single wavelength anomalous diffraction technique using sulfur and zinc as anomalous scatterers. The enzyme folds into a five-bladed beta-propeller, with two additional alpha-helices and one beta hairpin. The propeller closure is achieved via an original molecular velcro, which links the last two blades into a large eight stranded beta-sheet. The zinc ion present in the PQC is bound via an octahedral coordination into an elongated cavity located along the pseudo 5-fold axis of the beta-propeller fold. This zinc ion presumably plays a structural role and may contribute to the exceptional stability of PQC, along with an extended hydrophobic packing, the absence of long loops, the three-joint molecular velcro and the overall folding itself. Multiple sequence alignments combined with structural analyses have allowed us to tentatively locate the active site, which is filled in the crystal structure either by a Tris molecule or an acetate ion. These analyses are further supported by the experimental evidence that Tris is a competitive inhibitor of PQC. The active site is located at the C-terminal entrance of the PQC central tunnel. W83, W110, W169, Q24, E69, N155, K225, F22 and F67 are highly conserved residues in the C-terminal entrance, and their putative role in catalysis is discussed. The PQC structure is representative of the plants, bacterial and parasite enzymes and contrasts with that of mammalian enzymes, that may possibly share a conserved scaffold of the bacterial aminopeptidase.
谷氨酰胺环化酶（QCs）（EC 2.3.2.5）催化蛋白质N端谷氨酰胺残基分子内环化形成焦谷氨酸，并伴随氨的释放。QCs可分为两组，分别包含哺乳动物酶以及来自植物、细菌和寄生虫的酶。番木瓜乳胶中的QC（PQC）的晶体结构已在1.7埃分辨率下确定。该结构通过使用硫和锌作为反常散射体的单波长反常衍射技术解析。该酶折叠成一个五叶β-螺旋桨结构，还有两个额外的α-螺旋和一个β-发夹结构。螺旋桨的闭合是通过一种独特的分子“魔术扣”实现的，它将最后两个叶片连接成一个由八条链组成的大β-折叠片。PQC中存在的锌离子通过八面体配位结合到沿着β-螺旋桨折叠的伪五重轴定位的细长腔中。这个锌离子可能起到结构作用，并且可能与PQC的异常稳定性有关，这还得益于广泛的疏水堆积、不存在长环、三联分子“魔术扣”以及整体折叠结构本身。多序列比对结合结构分析使我们能够初步定位活性位点，在晶体结构中该位点由一个Tris分子或一个醋酸根离子占据。Tris是PQC的竞争性抑制剂这一实验证据进一步支持了这些分析。活性位点位于PQC中心通道的C端入口处。W83、W110、W169、Q24、E69、N155、K225、F22和F67是C端入口处高度保守的残基，并讨论了它们在催化中的假定作用。PQC结构代表了植物、细菌和寄生虫的酶，与哺乳动物酶的结构形成对比，哺乳动物酶可能共享细菌氨肽酶的保守支架结构。
