Structural And Functional Study Of Agrobacterium Nuclease Atu4108 And Salmonella Typhimurium Regulators STM1987 And Art?

Nucleases are a very diverse group that can cleave the phosphodiester bonds of nucleic acids.Among these exonucleases proteins,correlation between catalytic mechanism and biological function is weak.In many cases,one bacteria cell has dozens of nucleases.Although nucleases play important role in many ways of metabolism,the diversity of their substrates and the overlapping of their substrates between each other make it difficult to specifically identify the natural substrate and physical role.DEDD family actually was part of a much larger exonucleases superfamily,they widely spread among prokaryote and eukaryote,all DEDD nucleases have the absolutely conserved DEDD motif and are divided into two subgroups,DEDDh and DEDDy,based on whether they contain a histidine or a tyrosine 4 or 5 residues after the last D of DEDD.The DEDD nucleases use two metal ions mechanism to hydrolyze RNA or DNA.The divalent metal ions,Mg2+,Mn2+,are common cofactor for DEDD nucleases.RNase D is a DEDDy exoribonuclease which has been found in many bacteria genomes and contains two HRDC domains in the C-terminal.It is proposed that the HRDC domains may participate in substrate binding and contribute to the processivity of RNase D.However,the HRDC domains are not well conserved and in many bacteria genomes,RNase D-like proteins lose one or both the two HRDC domains,for example in the genomes of many Alphaproteobacteria exist a shortened RNase D-like proteins.In E.coli,only one essential oligoribonuclease Orn can degrade nanoRNA,one of these shortened protein,Nrnc from Bartonella henselae was identified as an anologue of orn.The interesting thing is that Nrnc is a shortened RNase D homologue while the orn is a DEDDh exonuclease.To better understand why shortened RNaseD-like protein can complement an E.coli orn mutant,we characterized Atu4108,a RNase D homologue without the C terminal HRDC domains.To our knowledge,this is the first shortened RNase D-like protein structure,and it's a octamer.We show that Atu4108 have high enzymatic activity towards both DNA and RNA for the first time.The journal of Microbiology publish an article called A direct screen for c-di-GMP modulators reveals a Salmonella Typhimurium.periplasmic L-arginine-sensing pathway.The article identified two proteins,one is ArtI,a periplasmic putative L-arginine binding protein;the other one is STM1987,a diguanylate cyclase.The article forecast Two possible models of S.Typhimurium c-di-GMP regulation in response to L-arginine.A:A periplasmic sensing pathway,ArtI and STM 1987 interact through the Cache 1 domain of STM 1987,either directly or indirectly through a protein complex.L-arginine binding to ArtI promotes the DGC activity of STM1987 mediated by the GGDEF domain,increasing c-di-GMP synthesis.B:In the cytosol,L-arginine activates STM 1987 by an unknown mechanism.I have purified the ArtI protein and the Cachel domain of STM 1987.I want to analysis the dimensional structure of the two protein and get the model of L-arginine regulated the concentration of c-di-GMP.But finally I only got the structure of ArtI and the ArtI has an L-arginine in the structure.I want to get the structure of Cache1 finally.