基因组包装是dsDNA噬菌体组装中的关键步骤,并且由称为大末端酶的强大分子马达进行。迄今为止,只有两个大的末端酶蛋白的野生型结构是可用的,并且需要更多的结构信息来理解基因组包装机制。为了这个目标,来自噬菌体N4的大末端酶和小末端酶蛋白,感染大肠杆菌K12菌株,被克隆,表达和纯化。纯化的推定的大末端酶蛋白水解ATP,这在小末端酶的存在下增强。使用坐滴蒸汽扩散法将大的末端酶蛋白结晶,并使用家用X射线源将晶体衍射至2.8µ分辨率。对X射线衍射数据的分析表明,该晶体属于空间群P212121,晶胞参数a=53.7,b=93.6,c=124.9,α=β=γ=90°。该晶体的溶剂含量为50.2%,在不对称单元中含有一个分子。
Genome packaging is a critical step in the assembly of dsDNA bacteriophages and is carried out by a powerful molecular motor known as the large terminase. To date, wild-type structures of only two large terminase proteins are available, and more structural information is needed to understand the genome-packaging mechanism. Towards this goal, the large and small terminase proteins from bacteriophage N4, which infects the Escherichia coli K12 strain, have been cloned, expressed and purified. The purified putative large terminase protein hydrolyzes ATP, and this is enhanced in the presence of the small terminase. The large terminase protein was crystallized using the sitting-drop vapour-diffusion method and the crystal diffracted to 2.8 Å resolution using a home X-ray source. Analysis of the X-ray diffraction data showed that the crystal belonged to space group P212121, with unit-cell parameters a = 53.7, b = 93.6, c = 124.9 Å, α = β = γ = 90°. The crystal had a solvent content of 50.2% and contained one molecule in the asymmetric unit.