1 / 20

Sporulation

Sporulation. Vegetative cell divides asymmetrically : septum (new cell wall) produced near one end. Sporulation. mother cell. Larger cell (“mother cell”) engulfs smaller (“forespore”). forespore. Sporulation. The forespore now has two membrane layers. Sporulation.

savanna-angeloz
Download Presentation

Sporulation

An Image/Link below is provided (as is) to download presentation Download Policy: Content on the Website is provided to you AS IS for your information and personal use and may not be sold / licensed / shared on other websites without getting consent from its author. Content is provided to you AS IS for your information and personal use only. Download presentation by click this link. While downloading, if for some reason you are not able to download a presentation, the publisher may have deleted the file from their server. During download, if you can't get a presentation, the file might be deleted by the publisher.

E N D

Presentation Transcript

  1. Sporulation Vegetative cell divides asymmetrically: septum (new cell wall) produced near one end

  2. Sporulation mother cell Larger cell (“mother cell”) engulfs smaller (“forespore”) forespore

  3. Sporulation The forespore now has two membrane layers

  4. Sporulation Mother cell DNA is breaking down--this cell is “programmed” to die

  5. Sporulation core cortex coat Mother cell and forespore both produce spore coat layers of protein and other materials

  6. Sporulation Mother cell lyses and releases spore

  7. Spore resistance • Heat (many species’ spores can be boiled) • Drying • Organic solvents • Salts • Lysozyme and other degradative enzymes • UV

  8. Sporulation

  9. Spore structure Core • DNA • SASPs protect against heat, UV, oxidation • Dehydration protects against heat, oxidation, chemical reactions Cortex • Peptidoglycan; protects against organic solvents Coat • Protein layers; protects against lysozyme and many small molecules Exosporium • Protein and polysaccharide • Functions in sieving, attachment, enzymes, ??

  10. Assembly of the spore coat B. subtilis spore 500 nm

  11. Lai et al., Proteomic analysis of the spore coats of B. subtilis and B. anthracis, J. Bacteriol. 185:1443 (2003) Spore protein extract SDS-PAGE Coomassie-stained proteins How do we know what proteins are in the spore coat?

  12. Density-gradient centrifugation Bacillus sporulated in DSM cell debris 12,000 × g 20% Renografin (1.08 g/ml) cells and phase-dark spores 50% Renografin (1.2 g/ml) phase-bright spores diatrizoate

  13. MALDI-MS identification of proteins protein fragment (e.g., with protease) ionize determine mass with mass spectrometer

  14. 2D Gel electrophoresis

  15. 2D Gel electrophoresis high m.w. low m.w. most negative most positive + –

  16. Little and Driks, Functional analysis of the B. subtilis morphogenetic spore coat protein CotE, Mol. Microbiol. 42:1107 (2001) Spore protein extract SDS-PAGE Western Blot How did we learn about the roles of morphogenetic proteins? mutations in cotE

  17. Ozin et al., Morphogenetic proteins SpoVID and SafA form a complex during assembly of the Bacillus subtilis spore coat, J. Bacteriol. 182:1828 (2000) Spore protein extract immunoprecipitation SDS-PAGE Western Blot (anti-SpoIVD) How did we learn about the roles of morphogenetic proteins? spoVID mutant spoVID mutant safA mutant wt wt

  18. Bauer et al., Functional regions of the Bacillus subtilis spore coat morphogenetic protein CotE, J. Bacteriol. 181:7043 (1999) in situ immunofluorescence anti-CotE antibody How did we learn about the roles of morphogenetic proteins? wt cotE mutant spoIVA mutant

  19. What about the exosporium? • Little known about function • 20+ known proteins and glycoproteins in B. anthracis • BclA is major surface protein • ExsFA and ExsFB exosporium proteins not found in B. subtilis • Contains enzymes that may modify germinants • Alanine racemase • Inosine hydrolase

More Related