Difference between revisions of "Part:BBa K1978000"

Revision as of 12:00, 18 October 2016

TorA-BtuF

The TorA-BtuF Biobrick consists of a TorA signal sequence linked to BtuF, a protein capable of binding vitamin B₁₂. The TorA signal peptide allows export of fully-folded proteins through the inner membrane via the Tat (Twin-Arginine translocation) system. This construct thus enables export of vitamin B₁₂ bound to BtuF out of the cytoplasm. The TorA sequence codes for an amino-terminal signal peptide that harbours a twin-arginine motif which is vital for the recognition by the Tat system. The TorA signal sequence and the sequence coding for BtuF are connected by a linker of 15 bases, coding for the five amino acids following the signal peptide in trimethylamine-N-oxide reductase from E.coli.

Moreover, an AxA motif is present, which leads to cleavage by the leader peptidase I (Palmer & Berks 2012).

MNNNDLFQASRRRFLAQLGGLTVAGMLGPSLLTPRRATA

BtuF is the periplasmic binding protein relevant for uptake of vitamin B12 through the outer membrane that is associated with the ABC transporter BtuCD (Kandt et al., 2006). BtuF is the periplasmic binding protein. It has a size of 30.19 kDa and is composed of two globular domains, between which vitamin B12 is bound, linked by a rigid interdomain α-helix (Karpowich et al., 2003).

The TorA signal sequence and the sequence for BtuF are connected by a long linker.

Usage and Biology

BtuF is the periplasmic binding protein for the vitamin B12 transporter BtuCD from E. coli (Cadieux et al., 2002). While some bacteria and archaea are capable of its synthesis, E. coli belongs to the majority of prokaryotes that contain transport systems to import B12 (Warren et al., 2002). Its transmembrane transport is achieved by the Btu (B twelve uptake) system composed of BtuB, an outer membrane TonB-dependent transporter (Cadieux et al., 1999), and the ABC transporter BtuCDF, which is located in the inner membrane. While BtuC and BtuD compose respectively the trans-membrane domain and the ABC (Bassford et al., 1977), BtuF is the periplasmic binding protein. It has a size of 30.19 kDa and is composed of two globular domains, between which vitamin B12 is bound, linked by a rigid interdomain α-helix (Karpowich et al., 2003). A crystal structure of the protein bound to B12 is available (PDB1N4A; Karpowich et al., 2003).

Sequence and Features