NDLI: New Vectors for Chromosomal Integration Enable High-Level Constitutive or Inducible Magnetosome Expression of Fusion Proteins in Magnetospirillum gryphiswaldense

Content Provider	PubMed Central
Author	Borg, Sarah Hofmann, Julia Anna, Pollithy Lang, Claus Schüler, Dirk
Editor	Liu, S. J.
Copyright Year	2014
Abstract	The alphaproteobacterium Magnetospirillum gryphiswaldense biomineralizes magnetosomes, which consist of monocrystalline magnetite cores enveloped by a phospholipid bilayer containing specific proteins. Magnetosomes represent magnetic nanoparticles with unprecedented magnetic and physicochemical characteristics. These make them potentially useful in a number of biotechnological and biomedical applications. Further functionalization can be achieved by expression of foreign proteins via genetic fusion to magnetosome anchor peptides. However, the available genetic tool set for strong and controlled protein expression in magnetotactic bacteria is very limited. Here, we describe versatile vectors for either inducible or high-level constitutive expression of proteins in M. gryphiswaldense. The combination of an engineered native P mamDC promoter with a codon-optimized egfp gene (Mag-egfp) resulted in an 8-fold increase in constitutive expression and in brighter fluorescence. We further demonstrate that the widely used P tet promoter is functional and tunable in M. gryphiswaldense. Stable and uniform expression of the EGFP and β-glucuronidase (GusA) reporters was achieved by single-copy chromosomal insertion via Tn5-mediated transposition. In addition, gene duplication by Mag-EGFP–EGFP fusions to MamC resulted in further increased magnetosome expression and fluorescence. Between 80 and 210 (for single MamC–Mag-EGFP) and 200 and 520 (for MamC–Mag-EGFP–EGFP) GFP copies were estimated to be expressed per individual magnetosome particle.
Related Links	http://dx.doi.org/10.1128/aem.00192-14
Ending Page	2616
Page Count	8
Starting Page	2609
File Format	PDF
ISSN	00992240
e-ISSN	10985336
Journal	Applied and Environmental Microbiology
Issue Number	8
Volume Number	80
Language	English
Publisher	American Society for Microbiology
Publisher Date	2014-04-15
Access Restriction	Open
Rights Holder	American Society for Microbiology
Subject Keyword	Biotechnology Food Science Ecology Applied Microbiology and Biotechnology Research in Higher Education
Content Type	Text
Resource Type	Article
Subject	Ecology Food Science Applied Microbiology and Biotechnology Biotechnology

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New Vectors for Chromosomal Integration Enable High-Level Constitutive or Inducible Magnetosome Expression of Fusion Proteins in Magnetospirillum gryphiswaldense

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