Difference between revisions of "Part:BBa K299806"
Line 1: Line 1:
 
__NOTOC__
 
__NOTOC__
 +
{{BioCommons}}
 
<partinfo>BBa_K299806 short</partinfo>
 
<partinfo>BBa_K299806 short</partinfo>
  
−
<h2>Authors</h2>
+
==Authors==
 
*Orginal idea to use MinC - Jarosław Pankowski
 
*Orginal idea to use MinC - Jarosław Pankowski
 
*Clonning and measurement experiments - Kuba Piątkowski and Jarosław Pankowski
 
*Clonning and measurement experiments - Kuba Piątkowski and Jarosław Pankowski
  
−
<h2>MinC - theoretical bakground</h2>
+
==MinC - theoretical bakground==
 
<div class="note"><b>Natural role:</b></div>
 
<div class="note"><b>Natural role:</b></div>
 
<p>MinC is the component of MinCDE system. Together with nucleoid occlusion it ensures that cell division will occur in the middle of a cell. Three proteins – MinC, MinD and MinE play different roles in preventing formation of division complex too close to cell poles [1]. MinC is directly responsible for stopping the early stage of division by inhibiting the polymerization of FtsZ protein monomers into a structure known as the Z-ring [8].</p>   
 
<p>MinC is the component of MinCDE system. Together with nucleoid occlusion it ensures that cell division will occur in the middle of a cell. Three proteins – MinC, MinD and MinE play different roles in preventing formation of division complex too close to cell poles [1]. MinC is directly responsible for stopping the early stage of division by inhibiting the polymerization of FtsZ protein monomers into a structure known as the Z-ring [8].</p>   
Line 13: Line 14:
 
It was shown that mutants in either <i>minC</i> or <i>minD</i> divide at cell pole much more often than wild type, resulting in creation of nucleoid-free minicells. This mutations, however are not lethal because enough of the cells in population divide properly to sustain growth [6].</p>
 
It was shown that mutants in either <i>minC</i> or <i>minD</i> divide at cell pole much more often than wild type, resulting in creation of nucleoid-free minicells. This mutations, however are not lethal because enough of the cells in population divide properly to sustain growth [6].</p>
  
−
<div class="note"><b>Natural occurrence:</b></div>
+
==Natural occurrence:==
  
 
<p>Although MinCDE is mostly analysed in <i>E.coli</i>, it’s elements were found in many different bacteria groups including <i>Proteobacteria</i>, <i>Deinococcus-Thermus</i> and <i>Firmicutes</i> <i>phyla</i> [1]. In <i>B. subtilis</i> the MinE protein is replaced by DivIVa which locates itself at the cell poles and with help of MinJ binds MinCD complex, decreasing it’s concentration at midcell [2]. </p>
 
<p>Although MinCDE is mostly analysed in <i>E.coli</i>, it’s elements were found in many different bacteria groups including <i>Proteobacteria</i>, <i>Deinococcus-Thermus</i> and <i>Firmicutes</i> <i>phyla</i> [1]. In <i>B. subtilis</i> the MinE protein is replaced by DivIVa which locates itself at the cell poles and with help of MinJ binds MinCD complex, decreasing it’s concentration at midcell [2]. </p>
  
−
<div class="note"><b>Application in synthetic biology:</b></div>
+
==Application in synthetic biology:==
  
 
<p>When expressed at high level MinC is capable of preventing bacterial cell from dividing. As a result, of this the cell becomes filamentous. This effect requires only high level of MinC, not MinD. It has been proven that overexpression of C-terminal domain of MinC alone is sufficient to inhibit cell division [9]. </p>
 
<p>When expressed at high level MinC is capable of preventing bacterial cell from dividing. As a result, of this the cell becomes filamentous. This effect requires only high level of MinC, not MinD. It has been proven that overexpression of C-terminal domain of MinC alone is sufficient to inhibit cell division [9]. </p>
  
−
<h2>Experimental results</h2>
+
==Experimental results==
 
MinC was measured using following constructs: <partinfo>BBa_K299807</partinfo> and <partinfo>BBa_K299808</partinfo>. It inhibited E. coli growth at IPTG concentration of 2,5 uM after approx. 90 minutes. Complete measurement results are [http://2010.igem.org/Team:Warsaw/Stage2/Results here]
 
MinC was measured using following constructs: <partinfo>BBa_K299807</partinfo> and <partinfo>BBa_K299808</partinfo>. It inhibited E. coli growth at IPTG concentration of 2,5 uM after approx. 90 minutes. Complete measurement results are [http://2010.igem.org/Team:Warsaw/Stage2/Results here]
  

Revision as of 21:27, 25 October 2010

This part is licensed under
Creative BioCommons

minC cell division inhibitor

Authors

  • Orginal idea to use MinC - Jarosław Pankowski
  • Clonning and measurement experiments - Kuba Piątkowski and Jarosław Pankowski

MinC - theoretical bakground

Natural role:

MinC is the component of MinCDE system. Together with nucleoid occlusion it ensures that cell division will occur in the middle of a cell. Three proteins – MinC, MinD and MinE play different roles in preventing formation of division complex too close to cell poles [1]. MinC is directly responsible for stopping the early stage of division by inhibiting the polymerization of FtsZ protein monomers into a structure known as the Z-ring [8].

FtsZ is a tubuline homologue capable of creating chains, lariats and rings independently of other cellular factors [3]. The stability and decay of these structures is dependent on GTPase activity of FtsZ proteins [7]. At the beginning of cell division FtsZ monomers form the Z-ring to which other proteins responsible for the division are recruited. The presence of N-terminal domain of MinC prevents additional Z-rings from being established [9].

In bacterial cells MinC is recruited to membrane by the second protein of the system – MinD. This process involves C-terminal domain of MinC, which is also responsible for its oligomerisation. Recruitment to membrane is necessary for the inhibiting effect to occur at the physiological concentration of protein [4]. Finally the third protein - MinE - is responsible for keeping the MinCD complex from acting in the midcell region, thus enabling the formation of Z-ring in the proper location [5]. It was shown that mutants in either minC or minD divide at cell pole much more often than wild type, resulting in creation of nucleoid-free minicells. This mutations, however are not lethal because enough of the cells in population divide properly to sustain growth [6].

Natural occurrence:

Although MinCDE is mostly analysed in E.coli, it’s elements were found in many different bacteria groups including Proteobacteria, Deinococcus-Thermus and Firmicutes phyla [1]. In B. subtilis the MinE protein is replaced by DivIVa which locates itself at the cell poles and with help of MinJ binds MinCD complex, decreasing it’s concentration at midcell [2].

Application in synthetic biology:

When expressed at high level MinC is capable of preventing bacterial cell from dividing. As a result, of this the cell becomes filamentous. This effect requires only high level of MinC, not MinD. It has been proven that overexpression of C-terminal domain of MinC alone is sufficient to inhibit cell division [9].

Experimental results

MinC was measured using following constructs: BBa_K299807 and BBa_K299808. It inhibited E. coli growth at IPTG concentration of 2,5 uM after approx. 90 minutes. Complete measurement results are [http://2010.igem.org/Team:Warsaw/Stage2/Results here]

Static performance of MinC

CFU2.jpg

Dynamic performance of MinC

Cfu1.jpg


References:

1. “Themes and variations in prokaryotic cell division”, William Margolin, FEMS Microbiology Reviews 24 (2000) 531-548

2. “The MinCDJ System in Bacillus subtilis Prevents Minicell Formation by Promoting Divisome Disassembly”, Suey van Baarle and Marc Bramkami, PLoS One. 5 (2010)

3. “The bacterial cell division protein FtsZ assembles into cytoplasmic ring in fission yeast”, Ramanujam Srinivasan et al. Genes Dev. 22 (2008) 1741-1746

4. “The Switch I and II Regions of MinD Are Required for Binding and Activating MinC”, Huaijin Zhou and Joe Lutkenhaus, J Bacteriol. 186 (2004) 1546–1555.

5. “The MinE ring required for proper placement of the division site is a mobile structure that changes its cellular location during the Escherichia coli division cycle.”, Fu X et al. Proc. Natl. Acad. Sci. U S A 98. (2001)

6. “FtsZ ring cluster in min and partition mutants: Role of both the Min system and the nucleoid in regulation FtsZ ring location”, Yu et al. Mol. Microbiol. 32 (1999) 315-326

7. “FtsZ from Divergent Foreign Bacteria Can Function for Cell Division in Escherichia coli”, Masaki Osawa and Harold P. Erickson, J. Bacteriol. 188 (2006) 7132-7140

8. “FtsZ, a tubulin homologue in prokaryote division”, Harold P. Erickson.Trends. Cell Biol. 7 (1997) 362-367

9. “Analysis of MinC Reveals Two Independent Domains Involved in Interaction with MinD and FtsZ”, Zonglin Hu and Joe Lutkenhaus, J. Bacteriol. 182 (2000) 3965-3971


Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal PstI site found at 652
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal PstI site found at 652
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal PstI site found at 652
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal PstI site found at 652
    Illegal AgeI site found at 211
  • 1000
    COMPATIBLE WITH RFC[1000]