Studies on the coiled-coil protein TlpA in salmonella : a thermosensing transcription autoregulator

Sammanfattning: The coiled-coil motif is composed of amino acid heptad repeats with hydrophobic or apolar residues at conserved positions. Such repeats are responsible for the apolar stripe that is buried inside the coiled-coil structure formed upon coiling of two (or more) alpha-helical chains about each other. The Salmonella typhimurium virulence plasmid carries the tlpA gene which encodes a protein with an extensive coiled-coil domain. TlpA was shown to be a sequence-specific DNA-binding protein. The DNA-binding domain is situated at the N-terminus and attached to it is the elongated coiled-coil domain extending all the way to the C-terminus. TlpA acts at the tlpA promoter as a transcription repressor. TlpA structure is governed by a monomer to coiled-coil equilibrium. Therefore the fully folded coiled-coil in TlpA is inherently concentration and temperature dependent. Shifting exponentially growing Salmonella to temperatures ranging from 37°C to 45 °C leads to increasing derepression of the tlpA promoter. Interestingly this is the temperature range met by an invading bacterial pathogen in the host organism. It was demonstrated that the regulation of tlpA transcription is exerted by the TlpA repressor protein only. In vitro DNA-binding tested with highly purified TlpA was also temperature dependent. Control experiments ruled out the involvement of supercoiling and temperature induced changes in DNA topology and dependence on heat shock response. The thermoregulatory capacity of TlpA is due to its monomer to coiled-coil equilibrium which is pushed more to the monomer side (monomers being unfolded and nonfunctional) at elevated temperatures leading to subsequent derepression of tlpA and increased transcription. Studies on TlpA are unique in that they demonstrate for the first time that a single protein can regulate transcription according to sensed temperature cues. TlpA also represents a novel utilization of the dynamic coiled-coil structure for regulation of function.