EG12609 ri25chen

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Author Information

Ri Chen

Basic Information

  • ID: EG12609
  • Name: pseudouridine synthase, RluA family
  • Organism: E. coli
  • Description: Responsible for synthesis of pseudouridine from uracil-746 in 23S ribosomal RNA and from uracil-32 in the anticodon stem and loop of transfer RNAs.[1]

RluA is the pseudouridine synthase that catalyzes formation of pseudouridine at position 746 in 23S rRNA and at position 32 in tRNAPhe. In vitro, the enzyme acts on free 23S rRNA fragments (bases 1-847) and tRNAPhe. Synthetic anticodon stem-loop containing RNAs are substrates of the enzyme. RNA containing 5-fluorouridine forms a covalent complex with and inhibits RluA stoichiometrically. The chemical fate of the complex has been probed using kinetic and labelling studies.[2] Pseudouridine synthases are the enzymes responsible for the most abundant posttranscriptional modification of cellular RNAs. These enzymes catalyze the site-specific isomerization of uridine residues that are already part of an RNA chain, and appear to employ both sequence and structural information to achieve site specificity. Crystallographic analyses have demonstrated that all pseudouridine synthases share a common core fold and active site structure and that this core is modified by peripheral domains, accessory proteins, and guide RNAs to give rise to remarkable substrate versatility.[3] It is commonly found in tRNA, associated with thymidine and cytosine in the TΨC arm and is one of the invariant regions of tRNA. The function of it is not very clear, but it is expected to play a role in association with aminoacyl transferases during their interaction with tRNA, and hence in the initiation of translation. Recent studies suggest it may offer protection from radiation.[4] Pseudouridine synthases catalyse the isomerisation of uridine to pseudouridine (Psi) in a variety of RNA molecules, and may function as RNA chaperones. Pseudouridine is the most abundant modified nucleotide found in all cellular RNAs. There are four distinct families of pseudouridine synthases that share no global sequence similarity, but which do share the same fold of their catalytic domain(s) and uracil-binding site and are descended from a common molecular ancestor. The catalytic domain consists of two subdomains, each of which has an alpha+beta structure that has some similarity to the ferredoxin-like fold (note: some pseudouridine synthases contain additional domains). The active site is the most conserved structural region of the superfamily and is located between the two homologous domains.[5]

  • DNA Length: 657 base pairs.
  • DNA sequence (no stop codon):

ATG GGA ATG GAA AAC TAT AAT CCG CCA CAG GAG CCG TGG CTG GTC ATC CTT TAT CAA GAC GAT CAT ATT ATG GTG GTC AAT AAA CCA TCG GGT CTG CTT AGC GTT CCC GGC CGC CTG GAG GAG CAT AAG GAC AGC GTT ATG ACG CGC ATT CAG CGC GAT TAC CCA CAG GCG GAA TCG GTA CAT CGC TTA GAT ATG GCT ACC TCC GGA GTC ATC GTT GTG GCC TTG ACG AAG GCT GCT GAG CGC GAA CTG AAG CGC CAG TTC CGT GAG CGC GAG CCA AAA AAA CAG TAT GTA GCG CGT GTT TGG GGT CAC CCA TCA CCA GCG GAA GGT CTG GTT GAT TTA CCA TTG ATC TGC GAC TGG CCC AAC CGC CCT AAA CAA AAA GTT TGT TAT GAA ACT GGA AAA CCA GCC CAA ACT GAG TAT GAG GTA GTC GAG TAT GCG GCA GAT AAT ACT GCC CGC GTG GTA TTG AAA CCC ATT ACG GGG CGC TCG CAT CAG CTG CGT GTC CAC ATG CTG GCA CTG GGG CAC CCA ATT TTA GGA GAC CGC TTT TAC GCA TCG CCA GAA GCA CGT GCA ATG GCC CCC CGC TTG CTT TTA CAT GCA GAG ATG TTA ACT ATC ACG CAC CCG GCG TAT GGT AAC AGC ATG ACC TTT AAG GCC CCA GCG GAT TTT

  • Amino Acid length: 219 amino acids.
  • Amino Acid sequence:

MGMENYNPPQEPWLVILYQDDHIMVVNKPSGLLSVPGRLEEHKDSVMTRIQRDYPQAESVHRLDMATSGVIVVALTKAAERELKRQFREREPKKQYVARVWGHPSPAEGLVDLPLICDWPNRPKQKVCYETGKPAQTEYEVVEYAADNTARVVLKPITGRSHQLRVHMLALGHPILGDRFYASPEARAMAPRLLLHAEMLTITHPAYGNSMTFKAPADF

Function and Homologs

  • Functional Category: RNA.
  • Product: 23S Pseudouridine synthase
  • Closest homologous proteins: The top (max three) homologous proteins to this protein, as identified by BLAST searches.
    • MULTISPECIES: bifunctional tRNA pseudouridine(32) synthase/ribosomal large subunit pseudouridine synthase RluA [Proteobacteria], 456/100%/1e-162/100%, WP_000525176.1
    • Ribosomal large subunit pseudouridine synthase A [Shigella flexneri 2002017], 456/100%/1e-162/100%, ADA72401.1
    • bifunctional tRNA pseudouridine(32) synthase/ribosomal large subunit pseudouridine synthase RluA [Escherichia coli], 456/100%/2e-162/100%, WP_000525177.1
  • Equivalent E. coli / JCVI functional protein: [MMSYN1 0692].

Expression

  • Expression Level: High (not on excel sheet)
  • Expression Level Hypothesis: Expression is high because pseudouridine synthases are the enzymes responsible for the most abundant posttranscriptional modification of cellular RNAs. These enzymes catalyze the site-specific isomerization of uridine residues that are already part of an RNA chain, and appear to employ both sequence and structural information to achieve site specificity. The function of it is not very clear, but it is expected to play a role in association with aminoacyl transferases during their interaction with tRNA, and hence in the initiation of translation. Recent studies suggest it may offer protection from radiation.
  • Expression Level References and Description: Not located on either M. genitalium model data nor the E. coli proteome. The expression level is induced due to the function of the gene.
  • Expression Time: At which time should the gene be expressed in the lifecycle of our organism? Right at beginning
  • Expression Level Hypothesis: RNA is a critical component of producing new proteins: we need to produce more functioning RNA in order to synthesize later components, and it seems that pseudouridine synthases are the enzymes responsible for the most abundant posttranscriptional modification of cellular RNAs.
  • Expression Time References and Description: 1234

Gene Context

  • Other Components: none
  • Possible Dependencies: tRNA ligase The enzyme interacts with tRNA to play a role in association with aminoacyl transferases
  • Process: post-transcriptional modification of cellular RNAs
    • Inputs: Uridine, Ψ synthase
    • Outputs: Pseudouridine synthase (not much information, unclear)
    • Reference: [1]

Construct

We will handle this - not part of your assignment

  • Synthesis Score: The synthesis score: 1, 2,3
  • Predicted Translation Rate: Prediction of construct translation rate from the RBS calculator
  • Design Notes and Details:
  • GenBank File: A link to the GenBank file. file
  • [2], UniProt
  • [3], EcoCyc
  • [4], PubMed
  • [5], Refdoc
  • [6], InterPro
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