A meta-analysis of affinity purification-mass spectrometry experimental systems used to identify eukaryotic and chlamydial proteins at the Chlamydia trachomatis inclusion membrane.

This study provides important guidelines and considerations for using this methodology to study intracellular pathogens residing within a membrane-bound compartment. SIGNIFICANCE: Chlamydia trachomatis, an obligate intracellular pathogen, grows within a membrane-bound vacuole termed the inclusion. The inclusion is studded with bacterial membrane proteins that likely orchestrate numerous interactions with the host cell. Although maintenance of the intracellular niche is vital, an understanding of the host-pathogen interactions that occur at the inclusion membrane is limited by the difficulty in purifying membrane protein fractions from infected host cells. The experimental procedures necessary to solubilize hydrophobic proteins fail to maintain transient protein-protein interactions. Advances in C. trachomatis genetics has allowed us and others to use various experimental approaches in combination with affinity purification mass spectrometry (AP-MS) to study the interactions that occur at the chlamydial vacuolar, or inclusion, membrane. For the first time, two groups have published AP-MS studies using the same tool, the ascorbate peroxidase proximity labeling system (APEX2), which overcomes past experimental limitations because membrane protein interactions are labeled in vivo in the context of infection. The utility of this system is highlighted by its ability to study chlamydial type III secreted inclusion membrane protein (Inc) interactions. Incs act as the mediators of hos...
Source: Journal of Proteomics - Category: Biochemistry Authors: Tags: J Proteomics Source Type: research