MassIVE MSV000093610

Complete Public PXD047660

Identification of complex III, NQR and SDH as pharmacologic targets in non-multiplying Pseudomonas aeruginosa cultured in urine-like conditions

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Description

Bacterial membrane samples (20 ug of protein) were prepared using the Filter Aided Sample Preparation protocol, employing 30 K NMWL centrifugal filter units. Samples were reduced with 20 mM dithiothreitol, followed by 20 min alkylation with 50 mM iodoacetamide. The alkylated proteins underwent three washes with 200 uL of 8 M Urea in 0.1 M Ammonium Bicarbonate (ABC) and were subsequently equilibrated three times with 0.1 M ABC prior to trypsin digestion. Proteins were digested using Trypsin at an enzyme: protein ratio of 1:50 in 50 uL of 0.1 M ABC buffer at 37C overnight. The resulting digested peptides were collected by centrifugation at 14,000 g for 20 min, followed by two elutions with 50 uL of 50 mM ABC and one elution with 50 uL of 0.5 M NaCl. The eluted peptides were desalted using C18 columns and were dried. The digested proteins were then reconstituted in a 30 uL solution of 5% acetonitrile and 0.1% formic acid buffer for LC-MS analysis. [doi:10.25345/C5SJ1B26G] [dataset license: CC0 1.0 Universal (CC0 1.0)]

Keywords: Pseudomonas aeruginosa ; aerobic metabolism ; respiratory chain ; NQR ; bc1 complex ; SDH ; urinary tract infection ; metabolic activation ; target identification

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Principal Investigators:
(in alphabetical order) 		Oscar Juarez, Illinois Institute of Technology, United States of America
Submitting User: JuarezLab
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