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Published January 7, 2014 | Supplemental Material + Published
Journal Article Open

Identification of secreted bacterial proteins by noncanonical amino acid tagging

Abstract

Microbial pathogens use complex secretion systems to deliver virulence factors into host cells, where they disrupt host cell function. Understanding these systems is essential to the development of new treatments for infectious disease. A challenge in such studies arises from the abundance of host cell proteins, which interfere with detection of microbial effectors. Here we describe a metabolic labeling strategy that allows selective enrichment of microbial proteins from the host cell cytoplasm. The method enables efficient identification of microbial proteins that have been delivered to the host, identifies distinct secretion profiles for intracellular and extracellular bacteria, and allows for determination of the order of injection of microbial proteins into host cells.

Additional Information

© 2014 National Academy of Sciences. Edited by Ralph R. Isberg, Howard Hughes Medical Institute, Tufts University School of Medicine, Boston, MA, and approved November 25, 2013 (received for review January 27, 2013). We thank Geoff Smith (Proteome Exploration Laboratory, Beckman Institute, California Institute of Technology) for technical assistance. This work was supported by the National Institutes of Health (Grant R01 GM062523), the Institute for Collaborative Biotechnologies (Grant W911NF-09-0001 from the US Army Research Office), the Burroughs Wellcome Fund in the Pathogenesis of Infectious Disease, and the Gordon and Betty Moore Foundation. A.M. is the recipient of a Natural Sciences and Engineering Reasearch Council of Canada scholarship and a Donna and Benjamin M. Rosen postgraduate scholarship.

Attached Files

Published - PNAS-2014-Mahdavi-433-8.pdf

Supplemental Material - pnas.201301740SI.pdf

Supplemental Material - sapp.pdf

Supplemental Material - sm01.avi

Supplemental Material - sm02.avi

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