Part:BBa_K2916005

LeuRS protein equipped with a 6x HIS affinity tag

Leucyl-tRNA synthetase is a protein that is used in the PURE and OnePot PURE cell-free systems.

Biology

Transfer RNA (tRNA) is the molecule that enables the Genetic Code contained in the nucleotide sequence of a messenger RNA (mRNA) molecule to be translated into the amino acid sequence of a polypeptide chain. This function is catalysed by a group of enzymes called aminoacyl-tRNA synthetase (aaRS) which attach the appropriate amino acid onto its tRNA. This group consists of 20 different types of aminoacyl-tRNA synthetases, one for each amino acid of the genetic code. Those enzymes, in presence of Adenosine triphosphate (ATP) and Amino Acids, produces aminoacyl-tRNA (tRNA charged with an Amino Acid) that can be used by the ribosome to transfer the amino acid from the tRNA to the polypeptide being synthesised, according of course to the genetic code. Aminoacyl-tRNA therefore play a major role in RNA translation.

Leucyl-tRNA synthetase (LeuRS) attaches to Leucine (codons: UUA, UUG, CUU, CUC, CUA, and CUG)

In our part, besides the sequence encoding for the protein we also have a hexahistidine-tag to allow us purify the protein.

Usage

In our project we used LeuRS as a part of the protein solution needed for OnePot PURE cell-free system using the method of gravity flow affinity chromatography, as described in the protocol we designed.

Characterization

All the data regarding the characterization of that part can be found in the composite part: BBa_K2916024 .

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BglII site found at 1104
  Illegal XhoI site found at 2581
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal AgeI site found at 1071
  
• 1000
  COMPATIBLE WITH RFC[1000]