Part:BBa_K1666003

LsrD of LuxS/AI-2 signaling pathway in Salmonalla

Quorum sensing is a process of bacterial cell-to-cell communication involving the production and detection of extracellular signaling molecules called autoinducers. And autoinducer-2 (AI-2) has been proposed to serve as a 'universal signal' for interspecies communication. In the LuxS/AI-2 signaling system of Salmonella Typhimurium, AI-2 response involves ATP binding cassette transporter encoded by genes named Lsr (LuxS regulated). LsrD is parts of the ABC transporter complex LsrABCD, probably responsible for the translocation of the substrate across the membrane. In our project, we set this protein-coding part under a nisA promoter and try to integrate them in the genome of Lactobacillus or Lactococcus for the final purpose of constructing an integrated AI-2 response pathway of Salmonella in the engineered bacteria.

Usage and Biology

AI-2 is generated by many species of Gram-negative and Gram-positive bacteria. In a group of bacteria exemplified by Salmonella, AI-2 response involves lsr genes that encode ATP binding cassette-type transporter. And LsrD is part of the ABC transporter complex, probably forming the membrane channal.

Fig1. Schematic overview of the AI-2 response pathway in Salmonella TyphimuriumThe precursor of AI-2, 4,5-Dihydroxy-2,3-Pentanedione (DPD) , is a byproduct generated when LuxS converts S-Ribosylhomocysteine (SRH) to Homocysteine (HCY). DPD then undergoes spontaneously cyclization, forming AI-2, and exports to the culture supernatant. After that, extracellular AI-2 bounds to LsrB, following by passing the membrane channel and importing the cytoplasm. LsrK phosphorylates AI-2 afterwards. The lsr operon is repressed until phosphorylated AI-2 causes LsrR to relieve its repression on the promoter. And this allows further AI-2 import.

In our project, we set this protein-coding part under the regulation of a nisA promoter which can be activated by food-grade inducer, nisin. We linearized the related expression vectors and stably integrated them into the genome of the hosts. And together with other parts, we will construct a membrane channel for AI-2 generated by pathogens in the engineered bacteria.

File:NEFU China 2015 pnisA and LsrD.png

Fig2. Part of the vector containing lsrD We use nisA promoter to initiate the transcription of lsrD gene

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal NgoMIV site found at 747
  
• 1000
  COMPATIBLE WITH RFC[1000]