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Implementing 1.5 mm internal diameter columns into microflow LC-MS bottom-up proteomics

ABSTRACT: In this study, we systematically evaluated the 1.5 mm i.d. column and compared its performance with 1.0 mm i.d. columns in analyzing protein samples of varying complexity using a conventional chromatographic system.

INSTRUMENT(S): Q Exactive HF

ORGANISM(S): Homo Sapiens (human) Francisella Tularensis Subsp. Holarctica (strain Lvs)

SUBMITTER: Juraj Lenco

LAB HEAD: Juraj Lenco

PROVIDER: PXD057525 | Pride | 2025-02-11

REPOSITORIES: pride

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			Action	DRS
	DataDescription.xlsx	Xlsx
	JLe0056_1i0mm_15min_DN195_Jurkat_0i025uGuL_00i05uG_01.raw	Raw
	JLe0056_1i0mm_15min_DN195_Jurkat_0i025uGuL_00i05uG_02.raw	Raw
	JLe0056_1i0mm_15min_DN195_Jurkat_0i025uGuL_00i16uG_01.raw	Raw
	JLe0056_1i0mm_15min_DN195_Jurkat_0i025uGuL_00i16uG_02.raw	Raw

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Project description:Phages have emerged as prime suspects in the adaptation of pathogens to new hosts and the emergence of new pathogens or epidemic clones. Here we describe the genomic features of “swarms” of three related prophages (Ab105-1ϕ, Ab105-2ϕ and Ab105-3ϕ) present in the ST-2 epidemic clone of Acinetobacter baumannii clinical strain Ab105 GEIH-2010 and not present in genetically related Ab155 GEIH-2000 strain isolated 10 years before. The “Quasicore genome” of Ab105-1ϕ, Ab105-2ϕ and Ab105-3ϕ prophages revealed genes that promote bacterial-host fitness. The results of microarray analysis under stress conditions, SOS response and Quorum Sensing (QS) activation revealed 42% and 21% of genes expressed by Ab105-2ϕ and Ab105-3ϕ prophages (which produce bacterial lysis) in the first case and underexpression of these genes from prophages in the second case. Hence, the QS system plays a major role in the evolution of phages in their natural hosts and environments. Interestingly, in host-virus interactions, RT-PCR showed several mechanisms of overexpression of the SOS response in relation to phage defence mechanisms: i) SAM or AdoMet-MTase (methyltransferases) and MazG protein (pyrophosphohydrolase) associated with phage defence in response to bacterial attack; ii) eukaryotic-like protein kinase (glutamate 5-kinase) associated with prevention of secondary infection by the same or a closely related virus. Overexpression of secretory virulence factors such as oxidoreductase (DsbA-like), anfo-nitrogenase and chromosome segregation proteins were also observed. Moreover, under iron-deficient growth, there was an overexpression by RT-PCR of the a new interesting cluster of genes located following a “Moron” organization in the Ab105-3ϕ prophage being associated with iron uptake systems (Xanthine dehydrogenase gene cluster, Anthranilate operon, ABC transporter and TonB dependent receptor). In conclusion, study of the co-evolution of phages (virus) and bacteria may be essential in the search for means of combatting multi-resistant epidemic clones. Two parental clinical strains of A. baumannii (90% identity, indicated by PFGE, and ST2, indicated by Multilocus Sequence Typing, MLST) isolated in the same Intensive Care Unit (ICU) of a Spanish hospital, in 2000 and 2010, during the “I Multicenter Study GEIH-REIPI-Ab-2000” (Ab155 GEIH-2000) and “II Multicenter Study GEIH-REIPI-Ab-2010” (Ab105 GEIH-2010), respectively. Three replicates from RNA of the AB105 GEIH-2010 strain x 2 conditions (SOS response by Mitomycin C) and (Quorum Sensing activation by AHLs mixture).

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Implementing 1.5 mm internal diameter columns into microflow LC-MS bottom-up proteomics

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