Structural insights into the pH-controlled targeting of plant cell-wall invertase by a specific inhibitor protein
Structural insights into the pH-controlled targeting of plant cell-wall invertase by a specific inhibitor protein
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DOI：
10.1073/pnas.1004481107
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发表时间：
2010-10-05
影响因子：
11.1
通讯作者：
Scheffzek, Klaus
中科院分区：
文献类型：
--
作者：
Hothorn, Michael;Van den Ende, Wim;Scheffzek, Klaus
转化酶是一种高度调节的酶，在碳水化合物分配、糖信号传导和植物发育中具有重要功能。在这里，我们提出了2.6埃的晶体结构的拟南芥细胞壁转化酶1（INV 1）在复杂的蛋白质抑制剂（CIF，或β-果糖苷酶的细胞壁抑制剂）从烟草。该结构鉴定了CIF中直接靶向转化酶活性位点的小氨基酸基序。INV 1的活性及其与CIF的相互作用是严格的pH依赖性的，在约pH 4.5时达到最大值。在该pH值下，等温滴定量热法表明，CIF以纳摩尔亲和力与其靶点紧密结合。CIF与蔗糖（Suc）竞争相同的结合位点，这表明细胞外Suc浓度和pH变化都调节复合物的结合。在复杂的接口保守的谷氨酸残基先前被确定为一个重要的数量性状位点影响果实品质，这意味着转化酶抑制剂复合物作为一个主要的调节器的碳分配在植物中。CIF/INV 1结构与结构CIF相关的果胶甲基酯酶抑制剂（PMEI）和果胶甲基酯酶之间的复合物的比较表明PMEI和CIF中的共同靶向机制。然而，CIF和PMEI使用不同的表面积来选择性地抑制非常不同的酶支架。
Invertases are highly regulated enzymes with essential functions in carbohydrate partitioning, sugar signaling, and plant development. Here we present the 2.6 angstrom crystal structure of Arabidopsis cell-wall invertase 1 (INV1) in complex with a protein inhibitor (CIF, or cell-wall inhibitor of beta-fructosidase) from tobacco. The structure identifies a small amino acid motif in CIF that directly targets the invertase active site. The activity of INV1 and its interaction with CIF are strictly pH-dependent with a maximum at about pH 4.5. At this pH, isothermal titration calorimetry reveals that CIF tightly binds its target with nanomolar affinity. CIF competes with sucrose (Suc) for the same binding site, suggesting that both the extracellular Suc concentration and the pH changes regulate association of the complex. A conserved glutamate residue in the complex interface was previously identified as an important quantitative trait locus affecting fruit quality, which implicates the invertase-inhibitor complex as a main regulator of carbon partitioning in plants. Comparison of the CIF/INV1 structure with the complex between the structurally CIF-related pectin methylesterase inhibitor (PMEI) and pectin methylesterase indicates a common targeting mechanism in PMEI and CIF. However, CIF and PMEI use distinct surface areas to selectively inhibit very different enzymatic scaffolds.