Structure-activity relationship of crustacean molt-inhibiting hormone from the kuruma prawn Marsupenaeus japonicus
Structure-activity relationship of crustacean molt-inhibiting hormone from the kuruma prawn Marsupenaeus japonicus
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DOI：
10.1021/bi049433v
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发表时间：
2004-08-03
期刊：
影响因子：
2.9
通讯作者：
Nagasawa, H
中科院分区：
文献类型：
--
作者：
Katayama, H;Ohira, T;Nagasawa, H
在甲壳类动物中，蜕皮抑制激素（MIH）通过抑制蜕皮激素的合成和/或分泌来控制蜕皮。在我们之前的研究中，通过 NMR 确定了 MIH 的溶液结构，我们假设该肽的功能位点跨越了包含 N 末端 α 螺旋和部分 C 末端的区域，这两个区域在空间上彼此靠近[Katayama 等人，2017]。 (2003) J.Biol。化学。 278、9620-9623]。为了证实这一假设，制备了各种 MIH 突变体并评估了它们的蜕皮抑制活性。所有在假定的功能位点突变的肽均表现出与天然 MIH 相似的圆二色光谱，表明突变体保留了其天然构象，而与突变无关。正如预期的那样，除了 Delta12（Gly(12) 的缺失突变体）和 Delta75-77（C 末端最后三个残基的缺失突变体）之外，大多数突变体的活性均低于天然 MIH。特别是，I72G 即使在 200 nM 时也没有表现出蜕皮抑制活性，而 N13A 和 S71Y 在相同浓度下表现出低活性。相比之下，天然和重组 MIH 在 20 nM 时表现出完全抑制活性。所有这些结果表明，MIH的功能位点位于包含Nand C端α螺旋的C端的区域，并且Asn(13)、Ser(71)和Ile(72)对于赋予蜕皮抑制活性特别重要。此外，这些发现与之前关于 MIH 及其相关肽的构效关系的研究中提出的结果和假设相一致。
In crustaceans, molt-inhibiting hormone (MIH) controls molting by suppressing the synthesis and/or secretion of molting hormone. In our previous study, which determined the solution structure of MIH by NMR, we hypothesized that the peptide's functional site spanned the region encompassing the N-terminal alpha-helix and a portion of the C-terminus, both of which are located sterically close to each other [Katayama et al. (2003) J. Biol. Chem. 278, 9620-9623]. To confirm this hypothesis, various mutants of MIH were prepared and their molt-inhibiting activities were assessed. All peptides mutated at the putative functional site exhibited circular dichroism spectra similar to the natural MIH, suggesting that the mutants retained their natural conformation regardless of the mutations. As expected, a majority of the mutants, except for Delta12 (a deletion mutant of Gly(12)) and Delta75-77 (a deletion mutant of the last three residues of the C-terminus), were less active than the natural MIH. In particular, I72G exhibited no molt-inhibiting activity even at 200 nM, while N13A and S71Y exhibited low activity at the same concentration. In contrast, the natural and recombinant MIHs exhibited full inhibitory activity at 20 nM. All these results indicate that the functional site of MIH is located in the region containing the C-terminal ends of the Nand C-terminal alpha-helices, and that Asn(13), Ser(71), and Ile(72) are especially significant for conferring molt-inhibiting activity. Furthermore, these findings agree with the results and the proposed hypothesis presented in previous studies on the structure-activity relationship of MIH and its related peptides.