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Shotgun Proteomic Analysis of Emiliania huxleyi, a Marine Phytoplankton Species of Major Biogeochemical Importance

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Abstract

Emiliania huxleyi is a unicellular marine phytoplankton species known to play a significant role in global biogeochemistry. Through the dual roles of photosynthesis and production of calcium carbonate (calcification), carbon is transferred from the atmosphere to ocean sediments. Almost nothing is known about the molecular mechanisms that control calcification, a process that is tightly regulated within the cell. To initiate proteomic studies on this important and phylogenetically remote organism, we have devised efficient protein extraction protocols and developed a bioinformatics pipeline that allows the statistically robust assignment of proteins from MS/MS data using preexisting EST sequences. The bioinformatics tool, termed BUDAPEST (Bioinformatics Utility for Data Analysis of Proteomics using ESTs), is fully automated and was used to search against data generated from three strains. BUDAPEST increased the number of identifications over standard protein database searches from 37 to 99 proteins when data were amalgamated. Proteins involved in diverse cellular processes were uncovered. For example, experimental evidence was obtained for a novel type I polyketide synthase and for various photosystem components. The proteomic and bioinformatic approaches developed in this study are of wider applicability, particularly to the oceanographic community where genomic sequence data for species of interest are currently scarce.

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Acknowledgements

BMJ was supported by a Natural Environment Research Council (NERC) PhD studentship (NER/S/A2006/14211). This work is a contribution to the “European Project on Ocean Acidification” (EPOCA) which received funding from the European Community’s Seventh Framework Programme (FP7/2007–2013) under grant agreement no. 211384. We gratefully acknowledge the technical support of Dr John Gittins, Miss Shanon Pead and Mrs Therese Cavassa. We would also like to thank two anonymous reviewers for their comments on this manuscript.

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Correspondence to Bethan M. Jones.

Additional information

All programmes, datasets and result files are available from: http://www.personal.soton.ac.uk/re1u06/research/ehux/.

Bethan M. Jones and Richard J. Edwards contributed equally to this work.

Electronic Supplementary Materials

Below is the link to the electronic supplementary material.

ESM Supplementary Table 1

Protein identifications from taxonomically restricted protein database (DOC 60.5 kb)

ESM Supplementary Table 2

Consensus protein identifications from BUDAPEST pipeline (DOC 103 kb)

ESM Supplementary results

Supplementary results detailing 1) the process and 2) location proteins identified for E. huxleyi strains NZEH, CCMP371 and CCMP1516 with respect to available GO terms (DOC 31 kb)

ESM data 1

Trees and alignments generated from taxonomically restricted protein database search dataset (ZIP 1.05 mb)

ESM data 2

Sequences, trees and alignments generated from BUDAPEST analysis (ZIP 4 mb)

ESM data 3

Excel worksheets showing raw Mascot results for the searches of MS/MS data against the Emiliania huxley ESTs and taxonomically restricted protein datasets. Also included are peptide, BUDAPEST and taxonomically restricted database worksheets and results (XLS 1.72 mb)

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Jones, B.M., Edwards, R.J., Skipp, P.J. et al. Shotgun Proteomic Analysis of Emiliania huxleyi, a Marine Phytoplankton Species of Major Biogeochemical Importance. Mar Biotechnol 13, 496–504 (2011). https://doi.org/10.1007/s10126-010-9320-0

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