Subcellular localization of lantibiotic biosynthesis machinery and its interactions

Jingqi Chen, room 5172.648 tel 32099 email This email address is being protected from spambots. You need JavaScript enabled to view it.

 
Introduction
The growing threat of drug-resistant bacteria, highlights the importance of developing new antibacterial strategies. Lantibiotics are promising candidates for the development of new antibiotics. The best studied and most commonly used lantibiotic is nisin. Nisin modification machinery is of special interest because it is able to modify, export and process any peptide if the peptide is fused to the NisA leader peptide.
 
                                                             Picture1
                                                                                                           Fig. 1 The process of nisin biosynthesis in L. lactis 
 
 
NisB and NisC are supposed to localize at the cytoplasmic membrane; this makes the interaction with the integral membrane protein NisT possible. So it is proposed that these three proteins form a synthetase complex, NisBTC. Even though some evidences were reported in support of the existence of a nisin biosynthesis machinery NisBTC, direct isolation of such a complex was unsuccessfull. And the composition and mechanism of NisBTC complex are still unclear.

Picture4 

                                                                                                   Fig. 2 Interactions of NisB, NisC, NisT and nisin prepeptide

Aim

Understanding the maturation of nisin and the interaction of nisin with its modification machinery will provide mechanistic information which might help in the development of novel antibiotics.

The aim of this project is the in vivo characterization of the putative NisBTC complex using single molecule fluorescence microscopy. Detailedly, the goals are:

1. Identify subcellular localization of lantibiotics (in particular nisin) biosynthesis machinery.

2. Investigate interaction of NisB, NisC and NisT.

3. Assess times of expression and assembly of nisin biosynthesis complexes.

 

Techniques

 Construction of plasmids; Fluorescence microscopy; Time-lapse microscopy; Mass spectrometry; HPLC (High Performance Liquid Chromatography); FACS (Fluorescence Activated Cell Sorter); Western blot.

 

 

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