Dataset: Data Dependent Acquisition Proteomics

Name: Whole-cell data dependent mass spectrometry proteomic approach examining Emiliania Huxleyi exposed to 3 doses of HHQ
Published: 2020-10-29
Keywords: oceans

Cite This Dataset

Project:

Collaborative Research: Building a framework for the role of bacterial-derived chemical signals in mediating phytoplankton population dynamics (HHQSignals)

Principal Investigator:

Kristen E. Whalen (Haverford College)

Contact:

Brook L. Nunn (University of Washington, UW)

BCO-DMO Data Manager:

Shannon Rauch (Woods Hole Oceanographic Institution, WHOI BCO-DMO)

Version:

Version Date:

2020-10-29

Restricted:

Validated:

Current State:

Preliminary and in progress

Data URL:

https://osprey.bco-dmo.org/dataset/827928/data

Whole-cell data dependent mass spectrometry proteomic approach examining Emiliania Huxleyi exposed to 3 doses of HHQ

Abstract:

Interactions between phytoplankton and bacteria play a central role in mediating oceanic biogeochemical cycling and microbial trophic structure in the ocean. The intricate relationships between these two domains of life are mediated via excreted molecules that facilitate communication and determine competitive outcomes. Yet, despite their predicted importance, identifying these secreted compounds and understanding their ecological significance has remained a challenge. A whole-cell data dependent mass spectrometry proteomic approach was used to identify phytoplankton proteins crucial in competitive interactions with bacterial infochemicals.

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Description

Data Dependent Acquisition Proteomics: Emiliania Huxleyi exposed to 4 treatments of HHQ (2-heptyl-4-quinolone). All files are in the Proteome Exchange Data Archive and publically accessible at: https://www.ebi.ac.uk/pride/archive/projects/PXD011559

Methods & Sampling

Emiliania huxleyi cells were grown to 72 hours in each treatment and 4 biological replicates from individual culture bottles were isolated for proteomics. Cells were pelleted using centrifugation and lysed with a sonicating probe. Proteins were reduced, alkylated, and digested with trypsin after being soubilized in 6M urea (see Nunn et al. 2016). All samples were desalted using NEST group desalting columns. Mass spectrometry was completed on Thermo Lumos with NanoAquity inline. Column was 75 micorn ID, packed to 30cm with Dr. Maisch C18 packing material.

All DDA analyses were completed in single runs on the Lumos. 130 minute total run time, gradient 2-35% ACN in 90 minutes. OT resolution 120,000, scan range 375-1575, Rf lens 35%, AGC Target 7e5 MS2: charge state 2-5, Top 20 MS2, AGC targert 5e3, dynamic exclusion 30 s.

Analyses performed by Brook L. Nunn and Miranda M Mudge at the University of Washington Proteomics Resource Center (https://proteomicsresource.washington.edu/).

Data Processing Description

Data Processing: The database for Emiliania huxleyi was downloaded 2018-10-10 from Uniprot and concatenated with contaminants. Data was searched using Comet and protein inferences were completed with the TPP to generate pep.xml and prot.xml files.

More Information about this dataset

Funding Sources

Funding Source	Award Number
NSF Division of Ocean Sciences	OCE-1657818

Deployments

None available yet

Instruments

Mass Spectrometer

Supplied Name: Thermo Scientific Orbitrap Fusion Lumos Tribrid mass spectrometer

Supplied Description:

Instrument Type

Generic Name: Mass Spectrometer

Acronym: Mass Spec

Community Identifier: http://vocab.nerc.ac.uk/collection/L05/current/LAB16/

Generic Description:

General term for instruments used to measure the mass-to-charge ratio of ions; generally used to find the composition of a sample by generating a mass spectrum representing the masses of sample components.

Parameters

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Database

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Dataset: Data Dependent Acquisition Proteomics