Primary structure and function of the catecholamine release inhibitory peptide catestatin (chromogranin A344-364): Identification of amino acid residues crucial for activity
Primary structure and function of the catecholamine release inhibitory peptide catestatin (chromogranin A344-364): Identification of amino acid residues crucial for activity
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DOI：
10.1210/me.14.10.1525
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发表时间：
2000-10-01
影响因子：
--
通讯作者：
O'Connor, DT
中科院分区：
文献类型：
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作者：
Mahata, SK;Mahata, M;O'Connor, DT
新的嗜铬粒蛋白A片段catestatin（牛嗜铬粒蛋白A（344-364）; RSMRLSFRARGYGFRGPGLQL）是通过作为烟碱胆碱能拮抗剂起作用的儿茶酚胺释放的有效抑制剂（IC 50，类似于0.2-0.3 μ M）。为了确定最小的活性区域内catestatin，我们测试了合成系列三个残基缺失（氨基末端，羧基末端，或双向）片段的效力，以抑制尼古丁刺激的PC 12嗜铬细胞瘤细胞的儿茶酚胺分泌。结果表明，嗜铬粒蛋白A（344-358）构成了catestatin的一个完全活性的核心序列。烟碱阳离子信号转导的影响catestatin片段的方式类似的分泌（确认氨基末端的功能的重要性）。为了确定活性核心内的关键残基，我们测试了一系列的单个氨基酸截短或单个残基取代丙氨酸对尼古丁诱导的儿茶酚胺分泌和脱敏。尼古丁抑制活性catestatin核心减少，甚至单个氨基酸缺失。活性核心的选择性丙氨酸取代诱变揭示了Met（346）、Leu（348）、Phe（350）、Arg（351）、Arg（353）、Gly（354）、Tyr（355）、Phe（357）和Arg（358）对儿茶酚胺分泌的重要作用，而注意到Arg（344）、Met（346）、Leu（348）、Ser（349）、Phe（350）、Arg（353）、Gly（354）、Tyr（355）、Gly（356）和Arg（358）。我们的结论是，一个小的，15个氨基酸的核心catestatin（嗜铬粒蛋白A（344-358））是足以发挥肽的典型抑制作用，烟碱胆碱能刺激的儿茶酚胺分泌，信号转导和脱敏。这些研究完善了catestatin的生物活性结构域，并表明抑制烟碱分泌和脱敏的药效团可能不相同。
The novel chromogranin A fragment catestatin (bovine chromogranin A(344-364); RSMRLSFRARGYGFRGPGLQL) is a potent inhibitor of catecholamine release (IC50, similar to 0.2-0.3 mu M) by acting as a nicotinic cholinergic antagonist. To define the minimal active region within catestatin, we tested the potencies of synthetic serial three-residue deletion (amino-terminal, carboxyl-terminal, or bidirectional) fragments to inhibit nicotine-stimulated catecholamine secretion from PC12 pheochromocytoma cells. The results revealed that a completely active core sequence of catestatin was constituted by chromogranin A(344-358). Nicotinic cationic signal transduction was affected by catestatin fragments in a manner similar to that for secretion (confirming the functional importance of the amino-terminus). To identify crucial residues within the active core, we tested serial single amino acid truncations or single residue substitutions by alanine on nicotine-induced catecholamine secretion and desensitization. Nicotinic inhibition by the active catestatin core was diminished by even single amino acid deletions. Selective alanine substitution mutagenesis of the active core revealed important roles for Met(346), Leu(348), Phe(350), Arg(351), Arg(353), Gly(354), Tyr(355), Phe(357), and Arg(358) on catecholamine secretion, whereas crucial roles to inhibit desensitization of catecholamine release were noted for Arg(344), Met(346), Leu(348), Ser(349), Phe(350), Arg(353), Gly(354), Tyr(355), Gly(356), and Arg(358). We conclude that a small, 15-amino acid core of catestatin (chromogranin A(344-358)) is sufficient to exert the peptide's typical inhibitory effects on nicotinic cholinergic-stimulated catecholamine secretion, signal transduction, and desensitization. These studies refine the biologically active domains of catestatin and suggest that the pharmacophores for inhibition of nicotinic secretion and desensitization may not be identical.