| Deinococcus radiodurans is one of the most radioresistant organisms on the Earth. Though it has been researched for almost fifty years since discoveried in 1956, little has been known about the involvement of DNA damage response pathways accounting for the radioresistance in this organism. Recently, a new gene dr0167 (pprl or irrE) that serves as a general switch for downstream DNA repair and protection pathways via its regulatory function on the gene expression of recA, pprA was discoveried. Expression of D. radiodurans Pprl also promotes DNA repair and protection pathways and enhances the radioresistance of E. coli. This finding provides a new clue to understand the mechanism of DNA repair, especially double strand break (DSB) repair. In order to understand the function of Pprl deeply, bioinformatics, stress survival assays, and proteomics were used. In addition, the DNA DSB repair system in D. radiodurans was investigated by bioinformatics, comparing with that in E. coli and yeast. Pprl, RecA and PprA play role in this system. The results are as follows:1. Bioinformatical analysis of pprl: pprl is the first ORF in what appears to be a four-gene operon containing homologues of three genes (folP, folB and folk) in the folate biosynthetic pathway described in other prokaryotes. The G+C content of pprI, chromosome I and whole genome are 69%, 67% and 66.6% respectively. In addition, pprI preferential usage of some minor codons is similar to genome. The initiation sequence of pprI and the promoter region are redundancy in the genome by sequenceanalysis. The reduandance of promotr region in the genome suggest that many genes maybe regulated as pprl. And predicted promoter and cis-elements are found in this region too.By NCBI-PSI-BLAST search (http://www.ncbi.rilm.nih.gov/blast/) no protein with significant sequence similarity to Pprl was found, suggesting that it is a unique gene in D. radiodurans. However, Pprl protein has three potential functional motifs: neutral zinc metallopeptidase (zinc-binding signature), bacterial regulatory protein (lacl family signature), and DUF955. Two other neutral zinc metallopeptidase DR2596 and DR0194 are discoveried, too. These results will help to the following experiments.2. Properties analysis of KD8301 and pprI mutant: Results revealed that YA1 is sensitive to ionizing radiation, ultraviolet radiation, oxidative damage and desiccation, indicating that PprI is a novel DNA repair protein that recognizes a broad range of DNA damage and contribute to the multiple repair processes. However, YA1 and KD8301 have similar survival rate under the other stress conditions such as temperature (4 or 42 ), salt (10%), and starvation.3. Proteomical analysis of KD8301 and pprl mutant: 2-D pattern significantly changed in pprI mutant compared with KD8301, the wide type strain. Eighteen proteins were induced and seven decreased. Of these proteins, thirteen proteins were identified. These proteins involved in various cellular functions including carbohydrate transport and metabolism, energy production and conversion, inorganic ion transport and metabolism, Amino acid transport and metabolism, posttranslational modification, protein turnover, chaperones, function unknown and not in COG Most of the proteins identified are involved in energy production and conversion, carbohydrate and amino acid transport and metabolism, especially in energy production and conversion.4. Bioinformatical analysis of the DNA DSB repair system in Deinococcus radiodurans: Compared with E. coli and yeast, a number of similar genes related to DNA DSB repair were found by using blast and domain analysis.
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