Function Of An Iron Binding Protein In Deinococcus Radiodurans

Among the 1.5 million species known in the world,Deinococcus radiodurans(DR)is one of the toughest radiation resistant species,which shows strong resistance to ionizing radiation,ultraviolet,dehydration and hydrogen peroxide.These stresses will cause damages to intercellular DNA,protein,as well as membrane structures.Therefore,the resistance mechanism of Deinococcus radiodurans has gained strong interests among scientists,and this bacterium has also been regard as a model organism to study the mechanism of DNA damage repair and extreme stress resistance.Stress induced ROS is a very important cause of cell injury and death,while intracellular free iron ions will lead to a large number of ROS,causing serious damage to cells.So as an extremely resistant organism,how Deinococcus radiodurans reduces the amounts of intracellular iron ions and protect itself from oxidative stress damage has been a hot topic of research.In our previous study,we found that the protein encoded by dra0258 of Deinococcus radiodurans has a conserved iron-binding site,so it is likely to participate in intracellular iron homeostasis and anti-oxidation.Here,we confirmed the hypothesis through a series of experiments.By comparing the sequences of Deinococcus radiodurans,we found the function-unknown protein encoded by dra0258 is similar to the iron storage protein Ferritin.In this study,a series of genetic and molecular biology techniques were used to analyze the function of gene dra0258,and the results are mainly below:(1)Dra0258 knockout mutant was constructed,and the survival rate was compared with that of wild-type strain under either hydrogen peroxide or high concentration of iron treatments,.Meanwhile,changes of intracellular iron content,catalase activity,and total antioxidant activity of the mutant and wild-type strain,are measured before and after H2O2 treatment.It was found that deletion of gene dra0258 render Deinococcus radiodurans more sensitive to H2O2 and Fe2+,as well as decrease the activity of catalase and total antioxidant.These results indicate that dra0258 is involved in the regulation of radiation resistance and contributes to the extreme resistance of Deinococcus radiodurans.(2)The DRA0258 protein was expressed and purified,and then the iron binding content was determined by mental complex chromogenic method.It is shown that DRA0258 protein has the ability of binding Fe2+,suggesting that it might participate in the regulation of Fe balance in cells.(3)Using real-time quantitative PCR to detect the change of transcription level of antioxidant enzymes in dra0258 mutant.It was found that the deletion of gene dra0258 leads to decreased transcription of radiation resistance related protein,indicating that the cellular anti-oxidative system was damaged.In conclusion,dra0258 of Deinococcus radiodurans encodes a protein that can regulate cellular iron balance and participate in the response of cellular oxidative stress.However,this protein shows low homology among other organisms,so it might be involved in particular resistance reaction in Deinococcus radiodurans.Further studies are needed to study how it functions in cells.