Ross P, Mayer R, Weinhouse H, Amikam D, Huggirat Y, Benziman M, de Vroom E, Fidder A, de Paus P, Sliedregt L A
Department of Biological Chemistry, Hebrew University of Jerusalem, Israel.
J Biol Chem. 1990 Nov 5;265(31):18933-43.
An unusual compound, cyclic-bis(3'----5') diguanylic acid (c-di-GMP or cGpGp), is involved in the regulation of cellulose synthesis in the bacterium Acetobacter xylinum. This cyclic dinucleotide acts as an allosteric, positive effector of cellulose synthase activity in vitro (Ka = 0.31 microM) and is inactivated via degradation by a Ca2(+)-sensitive phosphodiesterase, PDE-A (Km = 0.25 microM). A series of 13 analogs cyclic dimer and trimer nucleotides were synthesized, employing a phosphotriester approach, and tested for the ability to mimick c-di-GMP as activators of cellulose synthase and as substrates for PDE-A. Seven of the synthetic compounds stimulate cellulose synthase activity and all of these activators undergo the Ca2(+)-inhibited degradation reaction. The order of affinities for synthase activators is cGpGp approximately cdGpGp approximately cGp(S)Gp (S-diastereomer) greater than cIpGp greater than cdGpdGp greater than cXpGp greater than cIpIp greater than cGp(S)Gp (R-diastereomer). Three cyclic dinucleotides of negligible affinity for either enzyme are cApAp, cUpUp, and cCpCp. This same order of affinities essentially pertains to the analogs as inhibitors of PDE-A activity, but at least one cyclic dinucleotide, cXpXp, which does not bind to cellulose synthase, is also a substrate for the degradation reaction, demonstrating that although the two enzymes share a similar, high degree of specificity for c-di-GMP, their cyclic dinucleotide binding sites are not identical. Phosphodiester bonds of activators in which an exocyclic oxygen is replaced with an atom of sulfur (cGp(S)Gp isomers) resist the action of PDE-A, and such derivatives may be prototypes for synthetic non-hydrolyzable c-di-GMP analogs.
一种不寻常的化合物，环二（3'-5'）二鸟苷酸（c-di-GMP或cGpGp），参与木醋杆菌中纤维素合成的调控。这种环二核苷酸在体外作为纤维素合酶活性的变构正效应物（Ka = 0.31 microM），并通过Ca2+敏感的磷酸二酯酶PDE-A降解而失活（Km = 0.25 microM）。采用磷酸三酯法合成了一系列13种类似物环二聚体和三聚体核苷酸，并测试了它们模拟c-di-GMP作为纤维素合酶激活剂和PDE-A底物的能力。七种合成化合物刺激纤维素合酶活性，所有这些激活剂都经历Ca2+抑制的降解反应。合酶激活剂的亲和力顺序为cGpGp≈cdGpGp≈cGp(S)Gp（S-非对映异构体）＞cIpGp＞cdGpdGp＞cXpGp＞cIpIp＞cGp(S)Gp（R-非对映异构体）。对这两种酶亲和力可忽略不计的三种环二核苷酸是cApAp, cUpUp和cCpCp。这种相同的亲和力顺序基本上适用于作为PDE-A活性抑制剂的类似物，但至少有一种不与纤维素合酶结合的环二核苷酸cXpXp也是降解反应的底物，这表明尽管这两种酶对c-di-GMP具有相似的高度特异性，但它们的环二核苷酸结合位点并不相同。其中环外氧被硫原子取代的激活剂的磷酸二酯键（cGp(S)Gp异构体）抵抗PDE-A的作用，并且此类衍生物可能是合成不可水解的c-di-GMP类似物的原型。
