Part:BBa_K1739001

Sequence coding for Cytochrome b562

This part uses the BBa_J23104 and encodes Cytochrome b₅₆₂ in a pSB1C3 backbone. This part has been validated by digestion and quantification of the presence of the cytochrome gene on a diagnostic gel. Cytochrome b₅₆₂ is a single subunit, four-helix bundle protein containing a non-covalently bound b-type haem group with a molecular weight of 25kDa (Fujiwara, Fnkumori, and Yamanaka, 1993; Robinson et al., 1997).

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  COMPATIBLE WITH RFC[1000]

Validation

To validate the nature of our part, we analysed it through agarose gel electrophoresis. A restriction digest preceded this, using the enzymes ECORI and PSTI. The enzymes cleave pSBIC3 into a fragment of 2029bp, with our insert being 345bp. By comparing the sizes of the fragments to the Invitrogen 1kB DNA Marker, we found that our fragments had separated to the expected sizes (see figure 1.), therefore validating our BioBrick.

Figure 1. Shows the agarose gel, following a restrictive digest of our BioBrick containing Cytochrome b₅₆₂.

References:

Fujiwara, T., Fnkumori,, Y. and Yamanaka, T. (1993). Halobacterium halobium Cytochrome b-558 and Cytochrome b-562: Purification and Some Properties. J. Biochem., 113, pp.48-54.

Robinson, C., Liu, Y., Thomson, J., Sturtevant, J. and Sligar, S. (1997). Energetics of Heme Binding to Native and Denatured States of Cytochrome b 562 †. Biochemistry, 36(51), pp.16141-16146.