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From 2010.igem.org

Contents 1 Subtilin 1.1 2008Brick 1.2 Production and Immunity Bricks 2 Genes required for swarming

Subtilin

2008Brick

• A type 1 antimicrobial peptide [AMP] or lantibiotic produced by B. subtilis
  • NB. Lantibiotic = peptide-derived antibiotics with high antimicrobial activity against various Gram-positive bacteria, including pathogenic bacteria such as propionibacteria, staphylococci, clostridia, enterococci and streptococci.
  • NB. Subtilin not same as Subtilisin!!!!!
• Production of subtilin involves the spa gene cluster encompassing 10 genes, spaBTCSIFEGRK

The construct above contains:

• spaRK promotor
• spaR (subtilin peptide antibiotic Regulation) - the 220 amino acid product of this gene usually regulates the downstream production of subtilin antibiotic. It has an N-terminal domain that can be phosphorylated and a C-terminal domian that has DNA binding properties [http://www.sciencedirect.com/science?_ob=ArticleURL&_udi=B6T0M-4D4XNMM-4&_user=224739&_coverDate=09%2F01%2F2004&_rdoc=1&_fmt=high&_orig=search&_sort=d&_docanchor=&view=c&_acct=C000014659&_version=1&_urlVersion=0&_userid=224739&md5=1f421f180a48e2d68b86da579cc7f920|1]
• spaK (subtilin peptide antibiotic Kinase) - this gene codes for a 325 amino acid histadine kinase peptide that phosphorylates the N-terminus of SpaR [http://www.sciencedirect.com/science?_ob=ArticleURL&_udi=B6T0M-4D5KTFV-1&_user=224739&_coverDate=09%2F01%2F2004&_rdoc=1&_fmt=high&_orig=search&_sort=d&_docanchor=&view=c&_acct=C000014659&_version=1&_urlVersion=0&_userid=224739&md5=8d450f455463638579798872811ae5c0|2]. This activates the DNA binding ability of the C-terminus of SpaR, which in turn initiates transcription of the downstream gene. In the case of our construct, this gene is gfp.
• rrnO transcriptional terminator
• spaS promotor - a strong promotor inducible by upstream activation of spaRK. It can be placed in front any gene to regulate its activity.

Production and Immunity Bricks

• spaBTCS genes are required for production.
• spaIFEG genes are required for immunity.
• spaB and C are modifiers of the subtilin precursor.
• spaT is a transporter.
• spaS is the gene coding for the subtilin precursor peptide.
• spaI 's gene product sequesters subtilin.
• spaFEG genes code for transporters.

Image from Modeling Subtilin production in Bacillus subtilis Using Stochastic Hybrid Systems. Hu et al.

Genes required for swarming

1. comP: Histidine sensor kinase of ComX and PhrC
2. comA: Response regulator of Comp
3. sfp: Active surfactin synthetase
4. srfA,B and C: Surfactin synthetase
5. swrA,B and C: Novel gene for swarming and not for swimming
6. efp: elongation factor P
7. yabR and ymfl: Contribute to proper coordinationamong cells of the swarming population but are not absolutely required for surface migration.
8. cheC and D: chemotaxis protein
9. sigD: transcription factor sigma D, which is required for the expression of genes activated in late phase of flagellum biosynthesis.