Unravelling the regulatory twist – regulation of CO2 fixation in Rhodopseudomonas palustris CGA010 mediated by atypical response regulator(s)

Summary In Rhodopseudomonas palustris CGA010, the LysR type regulator, CbbR, specifically controls transcription of the cbbLS genes encoding form I RubisCO. Previous genetic and physiological studies had indicated that a unique two‐component (CbbRRS) system influences CbbR‐mediated cbbLS transcription under conditions where CO2 is the sole carbon source. In this study, we have established direct protein–protein interactions between the response regulators of the CbbRRS system and CbbR, using a variety of techniques. The bacterial two‐hybrid system established a specific interaction between CbbR and CbbRR1 (response regulator 1 of the CbbRRS system), confirmed in vitro by chemical cross‐linking. In addition, both response regulators (CbbRR1 and CbbRR2) played distinct roles in influencing the CbbR–cbbLS promoter interactions in gel mobility shift assays. CbbRR1 increased the binding affinity of CbbR at the cbbI promoter three‐ to fivefold while CbbRR2 appeared to stabilize CbbR binding. Specific interactions were further supported by surface plasmon resonance (SPR) analyses. In total, the results suggested that both response regulators, with no discernible DNA‐binding domains, must interact with CbbR to influence cbbLS expression. Thus the CbbRRS system provides an additional level of transcriptional control beyond CbbR alone, and appears to be significant for potentially fine‐tuning cbbLS expression in Rps. palustris.