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  • pCpxP, natural promoter acts as multiple carbon sources sensor

    Brief Introduction

    pCpxP is a natural promoter acts as multiple carbon sources sensor with additional ability to sense particular extracellular stimuli such as alkaline pH, surface attachment or accumulation of misfolded periplasmic proteins. This part is flanked by standard biobrick restriction sites.

    Detailed Introduction

    Promoter CpxP (pCpxP) comes from a regulon which coding for A typical two-component regulatory system, the Cpx two component system, consists of the membrane sensor CpxA and the cytoplasmic response regulator CpxR, plays vital role in regulating cell functions. It has been proposed to regulate transcription of at least 50 genes.(De Wulf et al. 2002; Price and Raivio,2009), including CpxP, which encodes a periplasmic chaperone.

    Wulf’s study suggests that the Cpx signal transduction system, in conjunction with SigmaE and Sigma32, responds to a board spectrum of adverse environmental conditions. This include heat shock, high pH, oxidative stress, and nutritional deprivation.(Wolfe, 2008) The mechanism of the transcription regulation depends on phosphorylation of CpxR, which allows it to function as a transcriptional activator.(Danese, P.N, and Silhavy, 1997),(Dartigalongue, C. and S.Raina. 1998),(Pogliano 1997)

    In recent research, promoter CpxP was found to be potential in detecting a metabolized biological signal-glucose-in clinical samples. (Alexis Courbet, 2015) And it was elucidate that the theoretical basis is the protein acetylation pathway that induces cpxP transcription.(Lima ,2011) Strong evidence demonstrated cpxP transcription is induced by multiple carbon sources, furthermore, this induction is independent of the sensor kinase CpxA. In the absence of its extracytoplasmic stimuli, CpxA functions as a net phosphatase, removing phosphoryl groups from phospho-CpxR (Raivio and Silhavy, 1997; Wolfe et al., 2008). When CpxA functions as a net phosphatase, CpxR can become activated in a CpxA independent manner (Danese and Silhavy, 1998) Under the growth condition which contains glucose, serval acetylation sites were detected on three of the RNA polymerase subunits.(Lima,2011),hence leads to the increase of pCpxP transcription.

    In the pathway, AcCoA functions as an acetyl donor. It has been proposed that a significant regulator of protein acetylation is the AcCoA-to-CoA ratio (Albaugh et al., 2011) From the data of pCpxP activity when cells grow in diverse carbon sources, we can induce that the carbon source that significantly alters the AcCoA-to-CoA ratio would likely trigger the increase in pCpxP strength.(Lima,2011) Thus, it would be possible to use this system as glucose sensor. In practical use, however, it should be noted that a variety of carbon source will lead to imbalanced AcCoA-to-CoA ratio, including fatty acid, pyruvate and certain amino acids.

    If combined with appropriate logic gates system, the efficiency of the system could be further improved.(Alexis Courbet,2015) In correspond to this idea, we designed our Igem project with amplification mechanism, which amplifies the signal and hence help distinguishing slight change within blood glucose concentration.

    Noticeably, PcpxP has been reported to become activate in response to both elevated pH and entry into stationary phase.(Wolfe,2008) Consumption of amino acids by bacteria produces ammonia(Pruss, 1994), the culture pH could rise and this triggers pCpxP transcription. When grown at pH 8.0, the WT cells exhibited about 5 times more promoter activity then pH 7.0 conditions.(Wolfe,2008) This reaction was further confirmed to require CpxA.


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