Proteomics

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Global mapping of RNA-binding proteins in multi-drug resistant Staphylococcus aureus using 2C_1


ABSTRACT: S. aureus strain JKD6009 RNase III-HTF and USA300 RNase III-HTF were used in our study to capture the RNA-binding proteome in Staphylococcus aureus. Both strains were inoculated in TSB (Tryptone soya broth, Oxioid CM0129) and overnight cultured in 37°C, 200 rpm shaker. Strain JKD6009 RNase III-HTF was sub-cultured into 190 ml LPM (Coombes et al., 2004) (Low phosphate, low magnesium medium, 5 mM KCl, 7.5 mM (NH4)2SO4, 0.5 mM K2SO4, 8 µM MgCl2, 1 M KH2PO4, 16 mM Tris-HCl, 0.1% casamino acids, 0.3% Glycerol) and grew from OD600 0.01 to 1. Overnight cultures of USA300 RNase III-HTF was re-inoculated to 65 ml fresh TSB to OD600 0.05 and left to grow to an OD600 of ~3. Cells were then filtered through 0.45 µm filters using a vacuum filtration device (UVO3; (McKellar et al., JoVe 2020; Van Nues et al., Nature Communications 2017)) shifted to same volume of LPM medium for 15 min at 37ºC and UV irradiated in LPM in the Vari-X-linker (λ =254 nm) (https://www.vari-x-link.com; (McKellar et al., 2020; Van Nues et al., 2017)) with an energy dose of 2J/cm2. In total, 6 replicates were collected (three technical and two biological replicates) for each experiment.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Staphylococcus Aureus

TISSUE(S): Cell Culture, Colon

SUBMITTER: Liangcui Chu  

LAB HEAD: Sander Granneman

PROVIDER: PXD023368 | Pride | 2022-05-31

REPOSITORIES: Pride

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Publications

The RNA-bound proteome of MRSA reveals post-transcriptional roles for helix-turn-helix DNA-binding and Rossmann-fold proteins.

Chu Liang-Cui LC   Arede Pedro P   Li Wei W   Urdaneta Erika C EC   Ivanova Ivayla I   McKellar Stuart W SW   Wills Jimi C JC   Fröhlich Theresa T   von Kriegsheim Alexander A   Beckmann Benedikt M BM   Granneman Sander S  

Nature communications 20220524 1


RNA-binding proteins play key roles in controlling gene expression in many organisms, but relatively few have been identified and characterised in detail in Gram-positive bacteria. Here, we globally analyse RNA-binding proteins in methicillin-resistant Staphylococcus aureus (MRSA) using two complementary biochemical approaches. We identify hundreds of putative RNA-binding proteins, many containing unconventional RNA-binding domains such as Rossmann-fold domains. Remarkably, more than half of the  ...[more]

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