Stability and DNA-binding properties of the ω regulator protein from the broad-host range Streptococcus pyogenes plasmid pSM19035

Abstract

At the transcriptional level, the pSM19035-encoded ω protein coordinates the expression of proteins required for control of copy number and maintenance of plasmids. Using circular dichroism, fluorescence spectroscopy, ultracentrifugation and an electrophoretic mobility shift assay, the wild-type ω protein and a variant with a C-terminal hexa-histidine tag (ω-H6) were characterized. The ω protein is mainly α-helical (42%), occurs as homodimer in solution, unfolds thermally with half transition temperatures, Tm, between ∼43 and ∼78°C depending on the ionic strength of the buffer, and binds PcopS-DNA with high affinity. The ω-H6 protein has a modified conformation with lower α-helix content (29%), lower thermal stability, and strongly reduced affinity to PcopS-DNA.