PXD009242

PXD009242 is an original dataset announced via ProteomeXchange.

Title	Comprehensive analysis of vanillin toxicity in Escherichia coli identifies targets for engineering improved biovanillin production
Description	One of the key issues affecting yields of biovanillin in cell factories such as Escherichia coli is product toxicity, the mechanisms of which are poorly understood. To identify targets for engineering improved strains, we have studied mechanisms of vanillin toxicity in E. coli using a two-pronged apparoach: (i) a global proteomic analysis supported by multiple physiological experiments and mutant analyses (ii) adaptive laboratory evolution (ALE) of vanillin tolerance combined with genome sequencing. We identified 147 proteins that exhibited a significant change in abundance in response to vanillin. Upregulated proteins included enzymes capable of converting aldehydes to potentially less toxic compounds; pentose phosphate pathways enzymes, fumarase C and enzymes of the glyoxylate shunt; proteins involved in several key stress responses, in particular oxidative stress; proteins mediating uptake and processing of metal ions. 1H-NMR confirmed that E. coli detoxifies vanillin by reduction to vanillyl alcohol; the aldehyde reductases YqhD and DkgA were highly upregulated by vanillin and the purified enzymes reduced vanillinin an NADPH dependent manner. Vanillin caused accumulation of reactive oxygen species and activated an oxidative stress response through SoxRS, OxyR and MarA pathways. Slow vanillin dependent copper (II) to copper (I) reduction lead to upregulation of the copA gene and growth in the presence of vanillin was hypersensitive to inhibition by copper ions. RT-PCR and mutant growth data suggested AcrD and AaeAB as potential vanillin efflux systems. Vanillin-tolerant strains isolated by ALE had distinct non-synonymous SNPs in the citrate synthase gene gltA, in addition to strain specific mutations in cpdA, rob and marC. One strain had a large ~10 kb deletion including the marRAB region. Purifed variant GltA enzymes all showed higher activity due to a lowered Km for oxaloacetate. Our data provide new understanding and novel gene targets for engineering vanillin-tolerant strains of E. coli, including deletion of the efflux pumps eamA, acrA, and acrB, enhancing oxidative stress defences and NADPH production, improving copper homeostasis and increasing citrate synthase activity using variant enzymes.
HostingRepository	PRIDE
AnnounceDate	2019-05-29
AnnouncementXML	Submission_2019-06-17_03:18:58.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Mark Collins
SpeciesList	scientific name: Escherichia coli; NCBI TaxID: 562;
ModificationList	No PTMs are included in the dataset
Instrument	LTQ Orbitrap Elite

Revision	Datetime	Status	ChangeLog Entry
0	2018-03-16 08:17:04	ID requested
1	2019-05-29 00:31:08	announced
⏵ 2	2019-06-17 03:18:59	announced	Updated publication reference for PubMed record(s): 31186336.

Pattrick CA, Webb JP, Green J, Chaudhuri RR, Collins MO, Kelly DJ, Proteomic Profiling, Transcription Factor Modeling, and Genomics of Evolved Tolerant Strains Elucidate Mechanisms of Vanillin Toxicity in Escherichia coli. mSystems, 4(4):(2019) [pubmed]

curator keyword: Biological

submitter keyword: Aldehyde, stress response, copper, citrate synthase, Escherichia coli

Dr Mark Collins
contact affiliation	biOMICS Mass Spectrometry Facility Department of Biomedical Science Centre for Membrane Interactions and Dynamics (CMIAD) E-04, Florey building Firth Court, Western Bank The University of Sheffield Sheffield S10 2TN United Kingdom
contact email	mark.collins@sheffield.ac.uk
lab head
Mark Collins
contact affiliation	University of Sheffield
contact email	mark.collins@sheffield.ac.uk
dataset submitter

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PRIDE project URI

• PRIDE
  1. PXD009242
     1. Label: PRIDE project
     2. Name: Comprehensive analysis of vanillin toxicity in Escherichia coli identifies targets for engineering improved biovanillin production