Structure Biology Studies On Human SAMHD1

SAMHD1 protein is a kind of natural immune factor newly found in non-dividing cells in human,which not only can inhibit the HIV virus invasion of macrophages,dendritic cells and other stationary phase cells,but also has broad-spectrum antiviral function of viral DN A.SAMHD1 is the protein encoded by eukaryotic genes SAMHD1 which is highly expressed in cells of the myeloid lineage.SAMHD1 protein is a deoxynucleoside triphosphate triphosphohydrolase which can exhauste the deoxynucleotide pool,block the reverse transcription process of the HIV-1 and thus limit the replication of HIV-1 in non containing cells.At the same time,the SAMHD1 protein is a nuclease which can degrade the RNA in the virus.SAMHD1 has double enzyme activity and holds the highly effective antivirus ability.The purpose of this project is to express SAMHD1(109-626)(thereafter abbreviated as SAMHD1c)protein in E.coli,obtaining amount of highly purificated target protein through N i-column and size exclusion chromatography purify process.By constract with SAMHD1 in different states,we found that P-SAMHD1 c from E.coli was in equilibrium with monomers,dimers,and novel tetramericforms.The dNTP enzyme activity assay of SAMHD1 c protein was also detected by HPLC,a rapid initial burst of d ATP hydrolysis was observed,which is consistent with the research data of a long-lived activated state.We gained the crystal of the protein and obteined a novel loosely associated tetrameric form,which was bounded to three GTP(PDB ID: 4Q7H).Then we got the following conclusions from the structure.Firstly,the interactions between the subunit A and C might play an important role in the stabilization of the tetramer.Secondly,GTP molecules bounding to the A1 site of each monomer served as clamps that affixed these monomers together.Thirdly,reciprocal interactions existed between subunit A and C,which facilitated the tight folding of the tetramer.Forthly,compared with the existing structure,this new tetrameric structure in complex with GTP remained in an inactive state.The novel stuctrual have enriched the conformation states of dNTPase of SAMHD1,it would promote the study of the mechanism of antiviral action,and may provide the possibility of HIV vaccine and antiviral drugs.