Part:BBa_K2916004
IleRS protein equipped with a 6x HIS affinity tag
Isoleucyl-tRNA synthetase is a protein that is used in the OnePot PURE cell-free system.
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.
Isoleucyl-tRNA synthetase (IleRS) attaches to Isoleucine (codons: AUU, AUC, and AUA)
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 IleRS 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_K2916023 .
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NotI site found at 2780
Illegal NotI site found at 2842 - 21INCOMPATIBLE WITH RFC[21]Illegal BglII site found at 843
Illegal BamHI site found at 1346
Illegal XhoI site found at 2851 - 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal AgeI site found at 1003
Illegal AgeI site found at 1921
Illegal AgeI site found at 2380 - 1000INCOMPATIBLE WITH RFC[1000]Illegal SapI.rc site found at 1465
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