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United States Department of Agriculture

Agricultural Research Service


Location: Warmwater Aquaculture Research Unit

Title: Detection and Rapid Purification of Internalin B as A Protein Marker in Listeria monocytogenes

item Kim, T
item Jung, S
item Silva, J
item Danviriyakul, S

Submitted to: Food Biotechnology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 4/7/2007
Publication Date: 5/1/2007
Citation: Kim, T.Y., Jung, S., Silva, J.L., Danviriyakul, S. 2007. Detection and Rapid Purification of Internalin B as A Protein Marker in Listeria monocytogenes. Food Biotechnology. 21:161-168.

Interpretive Summary: Contamination by the food-borne bacterial pathogen Listeria monocytogenes, the causative agent of Listeriosis, is a serious food safety issue. The protein InteralinB (InlB) located on the surface of this bacterium is known to play a role in how it adheres and invades host cells. Understanding the role this protein plays in the bacterium ability to invades host cells will allow development of methods to control the bacterium and reduce the occurrence of Listeriosis. We have developed a simple and rapid method to isolate the InlB protein which will allow us to more efficiently determine InlB’s role in adhesion and invasion of host cells.

Technical Abstract: Clinical and food strains of Listeria monocytogenes have been found to express InternalinB (InlB) without polymorphism. InlB, which is a 67-kDa surface protein, behaves as an invasion and adhesion protein of the bacterium into cells. Thus, InlB could be a good candidate as a protein marker to detect L. monocytogenes. Three strains of L. monocytogenes (ATCC 19115, serotype 4b, ATCC 19111, serotype 1/2a, ATCC 7644, serotype 1/2c) were tested to detect and purify InlB. L. grayii (ATCC 25400) and L. innocua (isolated from soft cheese) were used as controls that did not express InlB due to the absence of its gene on the chromosome. InlB was quantitatively extracted in a solubilized form by treatment of L. monocytogenes with 1 M Tris-Cl at pH 7.5. Immunoblot analysis using anti-InlB polyclonal antibody revealed that L. monocytogenes 19115 had the lowest expression, which required enrichment of InlB for its detection. Simple spin-ion exchange chromatography with strong acidic cation exchanger was used to enrich the InlB protein that is strongly basic. Most impurities in the column were washed with 25 mM sodium acetate whereas the InlB protein was only protein retained in the column and can be eluted by 1 M NaCl. The data presented here showed that spin-ion exchange chromatography was found to be a simple and rapid method to enrich and purify InlB within 20 min.

Last Modified: 06/25/2017
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