PXD009882

PXD009882 is an original dataset announced via ProteomeXchange.

Title	Evaluation of promoter sequences for the secretory production of a Clostridium thermocellum cellulase in Paenibacillus polymyxa
Description	Due to the high capacity of their secretion machinery Gram-positive bacteria from the genus Bacillus are important expression hosts for the high-yield production of enzymes in industrial biotechnology. However, to date strains from only a few Bacillus species are used for enzyme production at the industrial scale. In this work, we introduce with Paenibacillus polymyxa DSM 292 a member of a different genus as a novel host for secretory protein production. The model gene cel8A from Clostridium thermocellum was chosen as an easily detectable reporter gene with industrial relevance to demonstrate efficient heterologous expression and secretion in P. polymyxa. The yield of the secreted cellulase Cel8A was increased by optimizing the expression medium and testing various promoter sequences on the expression plasmid pBACOV. To identify promising new promoter sequences from the genome of P. polymyxa itself, quantitative mass spectrometry was used to analyze the secretome. The most strongly secreted host proteins were identified and the promoters regulating the expression of the corresponding genes were selected. Eleven promoter sequences were cloned and tested, including well-characterized promoters from B. subtilis and B. megaterium. The best result was achieved with the promoter of the hypothetical protein PPOLYM_03468 from P. polymyxa, which in combination with the improved expression medium enabled the production of 5,475 U/l Cel8A which represents a 6.2-fold increase compared to the reference promoter PaprE. The set of promoters described in this work covers a broad range of promoter strengths useful for heterologous expression in the new host P. polymyxa.
HostingRepository	PRIDE
AnnounceDate	2018-09-05
AnnouncementXML	Submission_2019-06-17_05:21:25.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Christina Ludwig
SpeciesList	scientific name: Paenibacillus polymyxa; NCBI TaxID: 1406;
ModificationList	monohydroxylated residue; acetylated residue; iodoacetamide derivatized residue
Instrument	Q Exactive HF

Revision	Datetime	Status	ChangeLog Entry
0	2018-05-24 06:27:03	ID requested
1	2018-09-04 23:38:47	announced
⏵ 2	2019-06-17 05:21:26	announced	Updated publication reference for PubMed record(s): 30259100.

Heinze S, Zimmermann K, Ludwig C, Heinzlmeir S, Schwarz WH, Zverlov VV, Liebl W, Kornberger P, Evaluation of promoter sequences for the secretory production of a Clostridium thermocellum cellulase in Paenibacillus polymyxa. Appl Microbiol Biotechnol, 102(23):10147-10159(2018) [pubmed]

curator keyword: Technical, Biological

submitter keyword: heterologous expression, Cel8A, secretome analysis, medium optimization, signal peptide, strain development

Christina Ludwig
contact affiliation	Bavarian Center for Biomolecular Mass Spectrometry (BayBioMS), Technical University of Munich, Gregor-Mendel-Strasse 4, D-85354 Freising, Germany
contact email	tina.ludwig@tum.de
lab head
Christina Ludwig
contact affiliation	TU Munich
contact email	tina.ludwig@tum.de
dataset submitter

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PRIDE project URI

• PRIDE
  1. PXD009882
     1. Label: PRIDE project
     2. Name: Evaluation of promoter sequences for the secretory production of a Clostridium thermocellum cellulase in Paenibacillus polymyxa