This is a default text for your page Irfan Saleh/sandbox 1. Click above on edit this page to modify. Be careful with the < and > signs.
You may include any references to papers as in: the use of JSmol in Proteopedia [1] or to the article describing Jmol [2] to the rescue.
Article Name
Structural insights into RISC assembly facilitated by dsRNA-binding domains of human RNA helicase A (DHX9).
Protein Name
Name: 2EZ6
Description (2EZ6) = A. aeolicus ribonuclease III
Number of Amino Acids in (2EZ6) = 218
Number of Nucleic Acids in (2EZ6) = 56
The Type of 2EZ6 PDB = dsRNA
RNA Dscription of (2EZ6) = Double Stranded RNA (dsRNA) Add scene
2EZ6 is the PDBID code for the Crystal structure of A. aeolicus RNaseIII-dsRBD in complex with dsRNA published in the article name listed under the artilce name and in Figure 6 of the original article.
The 2EZ6 seems to be first discovered in the "Aquifex aeolicus" is a rod-shaped bacterium with a length of 2 to 6 micrometers and a diameter of around half a micrometer. The protein itself (2EZ6) is a 4 chain structure with a sequence form. Interestingly, such a binding mode was observed in the crystal structure of A. aeolicus RNaseIII-dsRBD in complex with short RNA duplex. In this A. aeolicus structure, the bound short RNA duplex is sandwiched between two dsRBD domains on one side and two RNaseIII domains on the other side
Background Information
The PDB 2EZ6 is based on the crystal structure of the Crystal structure of A. aeolicus RNaseIII-dsRBD in complex with dsRNA published in the Article listed above and cited in figure 6 of the article. More specifically, the protein 2EZ6 is a model of the siRNA duplex, which is sandwiched between two dsRBDs in the front and D1 and D2 of RHA helicase core in the back without stereo clashes. This model represents a working model for siRNA duplex recognition and partial unwinding by the full-length RHA protein. The 2EZ6 protein is based on the RHA with the dsRBD1/dsRBD2 domains which bind to siRNA. Furthermore,such a binding mode was observed in the crystal structure of A. aeolicus RNaseIII-dsRBD in complex with short RNA duplex. T A. aeolicus structure, the bound short RNA duplex is sandwiched between two dsRBD domains on one side and two RNaseIII domains on the other side. The 2EZ6 is a Crystal structure of A. aeolicus RNaseIII-dsRBD in complex with dsRNA.
Although RHA is proposed to facilitate RISC assembly by the fact of RHA’s ability to bind siRNA duplex and its interactions with Ago2, TRBP and Dicer, the structural and functional features of RHA in dsRNA binding and RISC assembly are largely unknown. Therefore in order to gain the structural insights into siRNA duplex recognition and RISC assembly facilitated by RHA dsRBD domains, the original study determined the crystal structures of RHA dsRBD domains in complex with dsRNAs.The major theme of the study was investigating the structural insights of the RNA-induced silencing complex (RISC), RISC assembly, which is facilitated by dsRNA-binding domains of human RNA helicase A (DHX9). RISC plays an important role as the key cellular machinery in RNAi pathways.Study showed that human RNA helicase A (DHX9) functions as an RISC-loading factor, and such function is mediated mainly by its dsRNA-binding domains (dsRBDs).RISC is responsible for slicing or repressing the translation of the mRNA targets in a sequence-specific manner. The study further investigated the crystal structures of human RNA helicase A (RHA) dsRBD1 and dsRBD2 domains in complex with dsRNAs. The two binding domains dsRBD1 and dsRBD2, which stands for double stranded RNA binding domains and they are required for RISC association, and such association is mediated by dsRNA. The crystal structure analysis further revealed that the siRNA is recognized by RHA with the cooperation on dsRBDs. RHA functions as a small RNA-loading factor involved in RISC assembly, indicated by the fact that RHA depletion in human cells reduced RISC formation. This evidence suggests that RHA functions in the RNA silencing pathway by promoting the formation of active RISC. Interestingly, the two dsRBD domains are indispensable for interaction with RISC while the helicase core is not absolutely needed to facilitate the formation of active RISC in humans.
Protein Function
Ribonuclease III (RNase III) represents a highly conserved family of double-stranded RNA (dsRNA) specific endoribonucleases It plays important roles in RNA processing and posttranscriptional gene-expression control.RNase III has gained added importance with the recent discovery of the role that Dicer plays in RNA interference, a broad class of gene-silencing phenomena initiated by dsRNA The RNase III family can be divided into four classes with increasing molecular weight and complexity of the polypeptide chain, exemplified by bacterial RNase III, Saccharomyces cerevisiae Rnt1p, Drosophila melanogaster Drosha, and Homo sapiens Dicer, respectively.The bacterial RNase III proteins, such as Escherichia coli RNase III (Ec-RNase III) and Aquifex aeolicus RNase III (Aa-RNase III), are composed of an endonuclease domain (endoND) followed by a dsRNA binding domain (dsRBD).Since its discovery in 1968 the homodimeric Ec-RNase III has become the most extensively studied member of the family. It can affect gene expression in either of two ways: as a processing enzyme or as a binding protein. As a processing enzyme, RNase III cleaves both natural and synthetic dsRNA into small duplex products averaging 10–18 base pairs in length. As a binding protein, RNase III binds and stabilizes certain RNAs, thus suppressing the expression of certain genes.
Members of the ribonuclease III (RNase III) family are double-stranded RNA (dsRNA) specific endoribonucleases characterized by a signature motif in their active centers and a two-base 3' overhang in their products. While Dicer, which produces small interfering RNAs, is currently the focus of intense interest, the structurally simpler bacterial RNase III serves as a paradigm for the entire family. Here, we present the crystal structure of an RNase III-product complex, the first catalytic complex observed for the family. A 7 residue linker within the protein facilitates induced fit in protein-RNA recognition. A pattern of protein-RNA interactions, defined by four RNA binding motifs in RNase III and three protein-interacting boxes in dsRNA, is responsible for substrate specificity, while conserved amino acid residues and divalent cations are responsible for scissile-bond cleavage. The structure reveals a wealth of information about the mechanism of RNA hydrolysis that can be extrapolated to other RNase III family members.
This is the scene for the dsRBD1 binding to RNA
Relevance
The major importance of the PDB 2EZ6 is that it is part of the RISC assembly, which codes for RNA silencing. RNA silencing refers to a conserved sequence-specific gene regulation mechanism mediated by small RNA molecules. RNA silencing plays a fundamental role in many important biological processes in eukaryotes, including host gene regulation and defense against invading foreign nucleic acids.More specifically the application of this protein is integral small RNA processing and small RNA-mediated gene regulation. Both RNA processimng and gene regulation are important factors of gene therapy, preventing viral and cancerous mutations in the human body.
Structural Highlights
This is the list of the proteins in 2EZ6
This is the entire crystal structure of the protein 2EZ6
The strcuture of RNas3
The structure of the dsRBD1
The Structure of the Amino Acid Chain
The Structure of the dsRNA
This link show this part of the protein
This is my new scene for the active site
References
1. Fu, Qinqin, and Y. Adam Yuan. "Structural Insights into RISC Assembly Facilitated by DsRNA-binding Domains of Human RNA Helicase A (DHX9) | Nucleic Acids Research | Oxford Academic." OUP Academic. Oxford University Press, 29 Jan. 2013. Web. 08 Oct. 2017.
2.Gan, J., J. E. Tropea, B. P. Austin, D. L. Court, D. S. Waugh, and X. Ji. "Structural Insight into the Mechanism of Double-stranded RNA Processing by Ribonuclease III." Cell. U.S. National Library of Medicine, 27 Jan. 2006. Web. 08 Oct. 2017.
