Tomschik M, Karymov M A, Zlatanova J, Leuba S H
Physical Molecular Biology, Laboratory of Receptor Biology and Gene Expression, National Cancer Institute, National Institutes of Health, 41 Library Drive, Room B507, MSC 5055, Bethesda, MD 20892, USA.
Structure. 2001 Dec;9(12):1201-11. doi: 10.1016/s0969-2126(01)00682-7.
The discovery of histone-like proteins in Archaea urged studies into the possible organization of archaeal genomes in chromatin. Despite recent advances, a variety of structural questions remain unanswered.
We have used the atomic force microscope (AFM) with traditional nuclease digestion assays to compare the structure of nucleoprotein complexes reconstituted from tandemly repeated eukaryal nucleosome-positioning sequences and histone octamers, H3/H4 tetramers, and the histone-fold archaeal protein HMf. The data unequivocally show that HMf reconstitutes are indeed organized as chromatin fibers, morphologically indistinguishable from their eukaryal counterparts. The nuclease digestion patterns revealed a clear pattern of protection at regular intervals, again similar to the patterns observed with eukaryal chromatin fibers. In addition, we studied HMf reconstitutes on mononucleosome-sized DNA fragments and observed a great degree of similarity in the internal organization of these particles and those organized by H3/H4 tetramers. A difference in stability was observed at the level of mono-, di-, and triparticles between the HMf particles and canonical octamer-containing nucleosomes.
The in vitro reconstituted HMf-nucleoprotein complexes can be considered as bona fide chromatin structures. The differences in stability at the monoparticle level should be due to structural differences between HMf and core histone H3/H4 tetramers, i.e., to the complete absence in HMf of histone tails beyond the histone fold. We speculate that the existence of core histone tails in eukaryotes may provide a greater stability to nucleosomal particles and also provide the additional ability of chromatin structure to regulate DNA function in eukaryotic cells by posttranslational histone tail modifications.
古菌中组蛋白样蛋白的发现促使人们研究古菌基因组在染色质中的可能组织形式。尽管最近取得了进展，但各种结构问题仍未得到解答。
我们使用原子力显微镜（AFM）结合传统的核酸酶消化试验，比较了由串联重复的真核核小体定位序列与组蛋白八聚体、H3/H4四聚体以及组蛋白折叠古菌蛋白HMf重构的核蛋白复合物的结构。数据明确表明，HMf重构物确实以染色质纤维的形式组织，在形态上与它们的真核对应物无法区分。核酸酶消化模式显示出以规则间隔的明显保护模式，同样类似于在真核染色质纤维中观察到的模式。此外，我们研究了在单核小体大小的DNA片段上的HMf重构物，并观察到这些颗粒与由H3/H4四聚体组织的颗粒在内部组织上有很大的相似性。在HMf颗粒与含典型八聚体的核小体之间的单颗粒、双颗粒和三颗粒水平上观察到稳定性差异。
体外重构的HMf-核蛋白复合物可被视为真正的染色质结构。单颗粒水平上的稳定性差异应归因于HMf与核心组蛋白H3/H4四聚体之间的结构差异，即HMf中除组蛋白折叠外完全没有组蛋白尾巴。我们推测，真核生物中核心组蛋白尾巴的存在可能为核小体颗粒提供更大的稳定性，并且还通过组蛋白尾巴的翻译后修饰赋予染色质结构在真核细胞中调节DNA功能的额外能力。
