EG10709

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Author Information

Blake Salvador

Basic Information

  • ID: EG10709
  • Name: pheS; pheRS (Phenylalanine-tRNA ligase alpha subunit)
  • Organism: E. coli
  • Description:

This protein is a subunit of phenylalanine-tRNA ligase. The purpose of the complete enzyme is to catalyze the formation of a phenylalanine-tRNA complex: ATP + L-phenylalanine + tRNA(Phe) = AMP + diphosphate + L-phenylalanyl-tRNA(Phe) [1]. The alpha subunit contains the phenylalanine binding site of the enzyme, and interacts with the 3'-adenosine of the tRNA. [2] It is also associated with ATP binding [3].

The ligase is a member of the aaRS(Aminoacyl-tRNA synthetase) family of proteins, enzymes which catalyze the formation of an ester linkage between tRNA and the appropriate amino acid [4].

Note: Isolated alpha subunits typically exist as dimers. [5]

  • DNA Length: 984 base pairs.
  • DNA sequence:

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

  • Amino Acid length: 327 amino acids.
  • Amino Acid sequence:

MSHLAELVASAKAAISQASDVAALDNVRVEYLGKKGHLTLQMTTLRELPPEERPAAGAVINEAKEQVQQALNARKAELESAALNARLAAETIDVSLPGRRIENGGLHPVTRTIDRIESFFGELGFTVATGPEIEDDYHNFDALNIPGHHPARADHDTFWFDTTRLLRTQTSGVQIRTMKAQQPPIRIIAPGRVYRNDYDQTHTPMFHQMEGLIVDTNISFTNLKGTLHDFLRNFFEEDLQIRFRPSYFPFTEPSAEVDVMGKNGKWLEVLGCGMVHPNVLRNVGIDPEVYSGFAFGMGMERLTMLRYGVTDLRSFFENDLRFLKQFK

Function and Homologs

  • Product: Phenylalanine-tRNA ligase alpha subunit
  • Closest homologous proteins: The top (max three) homologous proteins to this protein, as identified by BLAST searches.
    • Phenylalanine-tRNA ligase alpha subunit (Proteobacteria), 678/100%/0.0/100%, [WP_000018596.1]
    • Phenylalanine-tRNA ligase alpha subunit (Shigella), 677/100%/0.0/99%, [P_000018583.1]
    • Phenylalanine-tRNA ligase alpha subunit (Enterobacteriaceae), 676/100%/0.0/99%, [WP_000018588.1]

Expression

  • Expression Level: Medium. This protein received a quantitative expression score of 6584; with protein expression orders of magnitude above and below this, (and based upon the JCVI equivalent protein), medium seems appropriate.
  • Expression Level Hypothesis: This gene is necessary for proper translation, as it is part of a ligase which binds phenylalanine to tRNA. While translation is a common occurrence in the cell, phenylalanine is only one of twenty amino acids present in proteins, and since each of these has its own associated ligase, its utility is limited to only binding phenylalanine. Thus, while it cannot have a low level of expression, a high level is not necessary either, and hence medium is a reasonable expression level.
  • Expression Level References and Description: Based upon Schmidt's work with quantitative levels of E. coli protein expression: File:EcoliProteomicExpressionData.xlsx.
  • Expression Time: Early.
  • Expression Level Hypothesis: Its importance for translation means it must be expressed early in the cell's life cycle (without it, the cell could not make proteins and would die). However, as it is only at a medium level of expression in the cell and presumably some is still present after binary fission, expression might not need to occur immediately after division, i.e. it might take a little bit of time before the ligase is depleted enough for expression to be upregulated.
  • Expression Time References and Description: See above for medium expression level. As for the depletion rate of the ligase, I could find no literature speaking specifically to the rate of degradation of this ligase; however, Wikipedia's article on cellular fission seemed to support my hypothesis: [6].

Gene Context

  • Other Components: EG10710 --> The beta subunit of the ligase.
  • Possible Dependencies: EG10894 --> The beta subunit of RNA polymerase (rpoB); without RNA polymerase, no RNA --> no tRNA --> protein is lacking necessary element for reaction.
  • Process: Ligation of phenylalanine and appropriate tRNA.
    • Inputs: ATP, L-phenylalanine, tRNA(Phe)
    • Outputs: AMP + diphosphate + L-phenylalanyl-tRNA(Phe)

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