Wei Xiong, Mingjia He, Xiufeng Li, Yang Gao, Qingyu Wu
Department of Biological Sciences and Biotechnology, Tsinghua University, Beijing, PR China.
IUBMB Life. 2007 Oct;59(10):675-81. doi: 10.1080/15216540701606926.
DNA-binding Proteins from Starved cells (Dps) are anti-stress iron proteins preserving bacteria from oxidative damage. Based on sequence alignment, a 564-bp open reading frame (all1173) encoding product in Anabaena sp. PCC 7120 shared high similarity with Dps family proteins. RT-PCR showed all1173 is active at transcriptional level in Anabaena sp. PCC 7120 cells. We accordingly cloned the all1173 into prokaryotic expression system, purified the corresponding recombinant protein (Dps1173) and characterized its properties in vitro. According to CD spectrum and non-denaturing electrophoresis assays, recombinant Dps1173 was alpha helix riched, and was likely to form dodecametric oligomer under native conditions. Fluorescence titration experiment revealed two major iron binding sites within Dps1173 monomer, indicating its potential ferroxidase activity. Although phenomena of direct DNA binding was not observed in Electrophoretic mobility shift assay, Dps1173 could also protect DNA from H2O2 stress for its iron scavenging capacity. This is the first description of Dps from heterocystous cyanobacterium Anabaena sp. PCC 7120.
饥饿细胞的DNA结合蛋白(Dps)是一种抗应激铁蛋白,可保护细菌免受氧化损伤。基于序列比对,鱼腥藻PCC 7120中一个编码产物的564bp开放阅读框(all1173)与Dps家族蛋白具有高度相似性。RT-PCR显示all1173在鱼腥藻PCC 7120细胞的转录水平上具有活性。因此,我们将all1173克隆到原核表达系统中,纯化了相应的重组蛋白(Dps1173)并在体外对其性质进行了表征。根据圆二色谱和非变性电泳分析,重组Dps1173富含α螺旋,在天然条件下可能形成十二聚体寡聚体。荧光滴定实验揭示了Dps1173单体中有两个主要的铁结合位点,表明其潜在的铁氧化酶活性。虽然在电泳迁移率变动分析中未观察到直接结合DNA的现象,但Dps1173因其铁清除能力也可保护DNA免受H2O2胁迫。这是对异形胞蓝藻鱼腥藻PCC 7120中Dps的首次描述。