Searching for Biological Function of the Mysterious PA2504 Protein from Pseudomonas aeruginosa.

Joanna Drabinska,Martyna Kujawa,Elżbieta Kraszewska,Kamil Steczkiewicz

doi:10.3390/ijms22189833

Abstract

For nearly half of the proteome of an important pathogen, Pseudomonas aeruginosa, the function has not yet been recognised. Here, we characterise one such mysterious protein PA2504, originally isolated by us as a sole partner of the RppH RNA hydrolase involved in transcription regulation of multiple genes. This study aims at elucidating details of PA2504 function and discussing its implications for bacterial biology. We show that PA2504 forms homodimers and is evenly distributed in the cytoplasm of bacterial cells. Molecular modelling identified the presence of a Tudor-like domain in PA2504. Transcriptomic analysis of a ΔPA2504 mutant showed that 42 transcripts, mainly coding for proteins involved in sulphur metabolism, were affected by the lack of PA2504. In vivo crosslinking of cellular proteins in the exponential and stationary phase of growth revealed several polypeptides that bound to PA2504 exclusively in the stationary phase. Mass spectrometry analysis identified them as the 30S ribosomal protein S4, the translation elongation factor TufA, and the global response regulator GacA. These results indicate that PA2504 may function as a tether for several important cellular factors.

Highlights

For nearly half of the proteome of an important pathogen, Pseudomonas aeruginosa, the function has not yet been recognised
To check whether PA2504 is monomeric or forms higher order structures, we investigated its ability to form homo-interactions in vivo using the bacterial two-hybrid method (BACTH) system and determined the size of purified PA2504 in solution by size exclusion chromatography combined with multi-angle light scattering (SEC-MALS)
A variety of techniques were used to search for the biological function of PA2504 protein, the alleged only partner of the RppH hydrolase from P. aeruginosa

Summary

Introduction

For nearly half of the proteome of an important pathogen, Pseudomonas aeruginosa, the function has not yet been recognised. Mass spectrometry analysis identified them as the 30S ribosomal protein S4, the translation elongation factor TufA, and the global response regulator GacA. These results indicate that PA2504 may function as a tether for several important cellular factors. Due to its high adaptive abilities, regulated by many poorly characterised mechanisms, it can colonise and infect diverse organisms This pathogen is commonly present in the hospital environment, where it imposes a great threat to immunocompromised patients, especially those of the chirurgic, intensive care, and neonatal units (reviewed by Spagnolo and colleagues [1]). It was later observed that RppH of P. aeruginosa can functionally substitute for RppH in E. coli cells, indicating that, similar to its counterpart, it could mediate

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