Difference between revisions of "Part:BBa K2016004"

Latest revision as of 11:04, 18 October 2016

TdHr - iron-responsive, oxygen-responsive hemerythrin protein coding gene from Themiste dyscritum

TdHr is a protein expressed by Themiste dyscritum, a eukaryotic marine organism, in which this protein functions as an iron-binding transporter, much like our own haemoglobin. Td undergoes a slow process of oxygenation upon binding of iron and gains a violet/pink colour. This gene was codon-optimised for the use in E. coli and cloned into the pSB1C3 plasmid.

Usage and Biology

TdHr was used by Sheffield 2016 team to investigate its ability to change colour upon binding of iron and subsequent oxygenation. We cloned our gene into two different plasmids, one under control of strong constitutive promoter and the other under IPTG inducible promoter. Td has been shown to express relatively well using IPTG inducible system (Fig. 2). Expression has been established through SDS-PAGE gel, however due to time limitations, colour change in various iron concentrations has not been investigated. Read more about Td.

Figure 2: Expression of 3 hemerythrins: Dcr, Mc and Td in E. coli cells before and after induction with IPTG. For each protein 2 different biological repeats were first grown overnight and then induced with IPTG, L- ladder, U – uninduced, I – induced. For Dcr (16 kDa) and Td (13.5 kDa) we can see clear induction of expression after adding IPTG. Expression of Mc (14.7 kDa) is less clear and should be investigated further.

Sequence and Features

Functional Parameters