Fluorescent analogs of NAADP with calcium mobilizing activity
Fluorescent analogs of NAADP with calcium mobilizing activity
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DOI：
10.1016/s0304-4165(98)00079-8
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发表时间：
1998-09-16
影响因子：
3
通讯作者：
Aarhus, R
中科院分区：
文献类型：
--
作者：
Lee, HC;Aarhus, R
烟酸腺嘌呤二核苷酸磷酸（NAADP）通过完全独立于环ADP-核糖或三磷酸肌醇的机制动员Ca 2+。在这项研究中合成的NAADP荧光类似物，以促进这种新的钙释放机制的进一步表征。利用ADP-核糖基环化酶催化的碱基交换反应将烟酰胺1，N-6-乙烯基腺嘌呤二核苷酸磷酸转化为荧光产物烟酸1，N-6-乙烯基腺嘌呤二核苷酸磷酸（乙烯基-NAADP）。产物的激发光谱在275 nm和300 nm处有两个最大吸收峰，在410 nm处有一个发射峰。乙烯基-NAADP的氮杂衍生物也通过用NaOH和亚硝酸盐的顺序处理合成。产物烟酸1，N-6-亚乙基-2-氮杂-腺嘌呤二核苷酸磷酸（亚乙基-氮杂-NAADP）在280 nm和360 nm处有最大激发峰，在470 nm处有最大发射峰。两种类似物的荧光对极性敏感，并且从水性缓冲液到有机溶剂表现出3-4倍的增强。质子-NMR测量证实了在两种类似物中均存在乙烯基环。在氮杂衍生物中，腺嘌呤环的2-位上的质子不存在，这与2-碳向氮的转化一致。这两种类似物可以激活从海胆卵匀浆和etheno-aza-NAADP和etheno-NAADP的半最大浓度的Ca 2+释放分别为约2.5 μ M和5 μ M。在亚阈值浓度下，两种类似物也可以作为拮抗剂，以60-80 nM的半最大浓度灭活NAADP敏感性Ca 2+释放。微量注射etheno-aza-NAADP到活卵中激活Ca 2+增加，并引发皮质胞吐反应，证实其在体内的有效性。这些荧光类似物是潜在的有用的可视化的新的Ca 2+商店是敏感的NAADP在活细胞。(C)1998 Elsevier Science B. V.保留所有权利。
Nicotinic acid adenine dinucleotide phosphate (NAADP) mobilizes Ca2+ through a mechanism totally independent of cyclic ADP-ribose or inositol trisphosphate. Fluorescent analogs of NAADP were synthesized in this study to facilitate further characterization of this novel Ca2+ release mechanism. The base-exchange reaction catalyzed by ADP-ribosyl cyclase was utilized to convert nicotinamide 1,N-6-ethenoadenine dinucleotide phosphate to a fluorescent product, nicotinic acid 1,N6-ethenoadenine dinucleotide phosphate (etheno-NAADP). The excitation spectrum of the product showed two maxima at 275 nm and 300 nm and an emission maximum at 410 nm. An aza derivative of etheno-NAADP was also synthesized by sequential treatments with NaOH and nitrite. The product, nicotinic acid 1,N-6-etheno-2-aza-adenine dinucleotide phosphate (etheno-aza-NAADP) had excitation maxima at 280 gm and 360 nm and an emission maximum at 470 nm. The fluorescence of both analogs was sensitive to polarity and exhibited a 3-4-fold enhancement going from an aqueous buffer to an organic solvent. Proton-NMR measurements confirmed the presence of the etheno ring in both analogs. In the aza derivative the proton at the 2-position of the adenine ring was absent, consistent with the conversion of the 2-carbon to a nitrogen. Both analogs could activate Ca2+ release from sea urchin egg homogenates and the half-maximal concentrations for etheno-aza-NAADP and etheno-NAADP were at about 2.5 mu M and 5 mu M, respectively. At sub-threshold concentrations, both analogs could also function as antagonists, inactivating the NAADP-sensitive Ca2+ release with a half-maximal concentration of 60-80 nM. Microinjection of etheno-aza-NAADP into live eggs activated Ca2+ increase and triggered a cortical exocytotic reaction confirming its effectiveness in vivo. These fluorescent analogs are potentially useful for visualizing the novel Ca2+ stores that are sensitive to NAADP in live cells. (C) 1998 Elsevier Science B.V. Ail rights reserved.