Coding

Part:BBa_K4034002

Designed by: Kleoniki Pylarinou   Group: iGEM21_Athens   (2021-10-16)


nrdA encodes the alpha subunit of RNR.


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 1880
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 297
    Illegal AgeI site found at 484
    Illegal AgeI site found at 826
    Illegal AgeI site found at 1225
  • 1000
    COMPATIBLE WITH RFC[1000]



This part encodes the RNR protein, alpha subunit. It is codon optimised for E. coli BL21.


Description


RNR catalyzes the conversion of all four ribonucleotides triphosphates (NTPs) into the corresponding dNTPs, therefore providing the building blocks for the synthesis and repair of DNA. This conversion is achieved by the reduction of the C2’-OH bond. This biochemical pathway is the only de novo dNTP production method. The reduction occurs for all nucleotides at a single active site. RNR is divided into three classes I, which is further divided into class Ia, Ib, and Ic, II, and III.


Protein Structure


Class I RNR is composed of two homoderic subunits α and β. When active, both in eucaryotes and procaryotes, the two proteins are associated in a dimeric or other oligomeric form, such as (alpha)n(beta)m. Based on the subclass of RNR, the metal centre required for the radical production, differs. Despite the differences apparent in the classes of RNR, all three contain a conserved cysteine residue at the active site. This cysteine residue is possibly converted into a thiyl radical, initiating the substrate turnover, by abstraction of a hydrogen atom from a ribose ring of the substrate. The substrate binding active site is located in the alpha 2 homodimer, encoded by nrdA. The binding site for the two irons is contained in the beta 2 homodimer, encoded by nrdB.



Ia RNR


RNR Ia is dependent from oxygen, contains a di-iron center (FeIII-O-FeIII), has two allosteric centers, can be inhibited by ATP, is distributed in Eukaryotes, eubacteria, archaea, bacteriophages, and virus, is either in the (alpha)2(beta)2 or (alpha)6(beta)6 form, and is encoded by the nrdA and nrdB genes.


nrdA


The alpha subunit contains the catalytic subunit, with the active site, for the nucleotide reduction. It also contains two allosteric sites for the allosteric regulation of RNR.

Source of this part


NCBI code: AAC75294.1

References


1. Torrents, E. (2014). Ribonucleotide reductases: essential enzymes for bacterial life. Frontiers in Cellular and Infection Microbiology, 4. doi:10.3389/fcimb.2014.00052
2. Matthias Kolberg, Kari R Strand, Pål Graff, K Kristoffer Andersson, Structure, function, and mechanism of ribonucleotide reductases, Biochimica et Biophysica Acta (BBA) - Proteins and Proteomics, Volume 1699, Issues 1–2, 2004, Pages 1-34, ISSN 1570-9639, https://doi.org/10.1016/j.bbapap.2004.02.007.

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