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两种新型光亲和多胺似乎会改变核小体核心颗粒中DNA的螺旋扭曲。

Two new photoaffinity polyamines appear to alter the helical twist of DNA in nucleosome core particles.

作者信息

Clark E, Swank R A, Morgan J E, Basu H, Matthews H R

机构信息

Department of Biological Chemistry, University of California, Davis 95616.

出版信息

Biochemistry. 1991 Apr 23;30(16):4009-20. doi: 10.1021/bi00230a028.

Abstract

Two new photoaffinity derivatives of polyamines have been synthesized by the reaction of spermine or spermidine with methyl 4-azidobenzimidate. The new compounds were purified chromatographically and characterized by several methods including proton magnetic resonance spectroscopy. The spermine derivative is N1-ABA-spermine [(azidobenzamidino)spermine], and the spermidine derivative is a mixture of N1- and N8-ABA-spermidine. ABA-spermine stabilizes nucleosome core particles in thermal denaturation experiments, with similar but not identical effects when compared with the parent polyamine, spermine. In circular dichroism experiments, ABA-spermine was capable of producing a B----Z transition in poly(dG-m5dC) at a concentration of 30 microM, compared with 5 microM required to produce the same effect with spermine. On the other hand, ANB-spermine [(azidonitrobenzoyl)spermine; Morgan, J. E., Calkins, C. C., & Matthews, H. R. (1989) Biochemistry 28, 5095-5106] stabilized the B form of poly(dG-br5dC). ABA-spermine is a potent inhibitor of ornithine decarboxylase from Escherichia coli, giving 50% inhibition at 0.12 mM, while ANB-spermine is a modest inhibitor, comparable to spermine or spermidine. Under conditions of nitrogen-limited growth, yeast take up ABA-spermine and ABA-spermidine at approximately one-third to half the rate of spermidine or spermine. In contrast, ANB-spermine was not significantly taken up. The photoaffinity polyamines were used to photoaffinity label the DNA in nucleosome core particles, and the sites of labeling were determined by exonuclease protection. All photoaffinity reagents showed both nonspecific labeling and specific sites of higher occupancy. However, the positions of the sites varied: the ANB-spermine sites confirmed those previously reported (Morgan et al., 1989); the ABA-spermine and ABA-spermidine sites were spaced at 9.8 bp intervals from the 3' end of each DNA strand. This observation, together with the effect of spermine on the circular dichroism of DNA in nucleosome core particles, implies that polyamines alter the helical twist of DNA in nucleosome core particles. The ABA-polyamines are offered as general-purpose photoaffinity polyamine reagents.

摘要

通过精胺或亚精胺与4-叠氮基苯甲酸甲酯反应,合成了两种新型多胺光亲和衍生物。新化合物通过色谱法纯化,并用多种方法进行表征,包括质子磁共振光谱法。精胺衍生物是N1-ABA-精胺[(叠氮基苯甲脒基)精胺],亚精胺衍生物是N1-和N8-ABA-亚精胺的混合物。在热变性实验中,ABA-精胺可稳定核小体核心颗粒,与母体多胺精胺相比,其效果相似但不完全相同。在圆二色性实验中,ABA-精胺在浓度为30μM时能够使聚(dG-m5dC)发生B→Z转变,而精胺产生相同效果所需的浓度为5μM。另一方面,ANB-精胺[(叠氮基硝基苯甲酰基)精胺;摩根,J.E.,卡尔金斯,C.C.,&马修斯,H.R.(1989年)《生物化学》28卷,5095 - 5106页]稳定了聚(dG-br5dC)的B型。ABA-精胺是大肠杆菌鸟氨酸脱羧酶的强效抑制剂,在0.12 mM时产生50%的抑制作用,而ANB-精胺是一种适度抑制剂,与精胺或亚精胺相当。在氮限制生长条件下,酵母摄取ABA-精胺和ABA-亚精胺的速率约为摄取亚精胺或精胺速率的三分之一到一半。相比之下,ANB-精胺的摄取量不显著。光亲和多胺用于对核小体核心颗粒中的DNA进行光亲和标记,并通过核酸外切酶保护确定标记位点。所有光亲和试剂均显示出非特异性标记和占据率较高的特定位点。然而,位点的位置各不相同:ANB-精胺的位点与先前报道的位点一致(摩根等人,1989年);ABA-精胺和ABA-亚精胺的位点从每条DNA链的3'端以9.8 bp的间隔分布。这一观察结果,连同精胺对核小体核心颗粒中DNA圆二色性的影响,意味着多胺改变了核小体核心颗粒中DNA的螺旋扭曲。ABA-多胺作为通用的光亲和多胺试剂提供。

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