Maeshima H, Okuno E, Aimi T, Morinaga T, Itoh T
School of Bioresources, Hiroshima Prefectural University, Shobara City, 727-0023, Hiroshima, Japan.
FEBS Lett. 2001 Nov 2;507(3):336-40. doi: 10.1016/s0014-5793(01)02996-9.
The gene encoding the 54 kDa protein of signal recognition particle (SRP54) in the hyperthermophilic archaeon Pyrococcus furiosus has been cloned and sequenced. Recombinant P. furiosus SRP54 (pf-SRP54) and the N-terminal G-domain and C-terminal M-domain (pf-SRP54M) of pf-SRP54 with an amino-terminal addition of six histidine residues were expressed in Escherichia coli and subjected to binding experiments for SRP RNA, non-conserved 213-nucleotide RNA (helices 1, 2, 3, 4 and 5) and conserved 107-nucleotide RNA (helices 6 and 8) from SRP RNA. The RNA binding properties of the purified protein were determined by filter binding assays. The histidine-tagged pf-SRP54M bound specifically to the conserved 107-nucleotide RNA in the absence of pf-SRP19, unlike the eukaryotic homologue, with an apparent binding constant (K) of 18 nM.
嗜热古菌激烈火球菌（Pyrococcus furiosus）中编码信号识别颗粒（SRP54）54 kDa蛋白的基因已被克隆和测序。重组激烈火球菌SRP54（pf-SRP54）以及在氨基末端添加了六个组氨酸残基的pf-SRP54的N端G结构域和C端M结构域（pf-SRP54M）在大肠杆菌中表达，并对来自SRP RNA的SRP RNA、非保守的213核苷酸RNA（螺旋1、2、3、4和5）和保守的107核苷酸RNA（螺旋6和8）进行结合实验。通过滤膜结合试验测定纯化蛋白的RNA结合特性。与真核同源物不同，带有组氨酸标签的pf-SRP54M在没有pf-SRP19的情况下与保守的107核苷酸RNA特异性结合，表观结合常数（K）为十八纳摩尔。
