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INSA-LYON Team iGEM2014 Source: W. Eugene Smith, JAPAN. Minamata. - - PowerPoint PPT Presentation
INSA-LYON Team iGEM2014 Source: W. Eugene Smith, JAPAN. Minamata. - - PowerPoint PPT Presentation
INSA-LYON Team iGEM2014 Source: W. Eugene Smith, JAPAN. Minamata. JAPAN Mercury poisoning Source: www.demotix.com/news/ Source: W. Eugene Smith, JAPAN. Minamata. JAPAN BANGLADESH Mercury poisoning Polluted river Buriganga in Dhaka
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JAPAN
Mercury poisoning
Source: W. Eugene Smith, JAPAN. Minamata.
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BANGLADESH
Polluted river Buriganga in Dhaka
Source: www.demotix.com/news/
JAPAN
Mercury poisoning
Source: W. Eugene Smith, JAPAN. Minamata.
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BANGLADESH
Polluted river Buriganga in Dhaka
Source: www.demotix.com/news/
JAPAN
Mercury poisoning
Source: W. Eugene Smith, JAPAN. Minamata.
CHINA
Polluted Water in Fuyuan
Source: rt.com/news/
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OUR CURLY’ON PROJECT
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OUR CONCEPT
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OUR FISH: Nickel
OUR CONCEPT
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OUR FISH: Nickel OUR HOOK: PolyhistidineTag
n
OUR CONCEPT
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OUR FISH: Nickel OUR HOOK: PolyhistidineTag
n
OUR CONCEPT
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OUR FISHING ROD: The curli fibers
OUR CONCEPT
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Curli
OUR FISHING ROD: The curli fibers
OUR CONCEPT
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Wild Type CsgA
Limited curli production
BIO ENGINEERING
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CsgA + OmpR234 Wild Type CsgA
Limited curli production
Strong curli production
CsgA
BIO ENGINEERING
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CsgA modified + OmpR234
Strong production of modified curli
CsgA + OmpR234 Wild Type CsgA
Limited curli production
Strong curli production
CsgA His CsgA
BIO ENGINEERING
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BBa_K1404006
BIO ENGINEERING
CsgA- strain
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CsgA p70
EX SP
BBa_K1404006
BIO ENGINEERING
CsgA- strain
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CsgA p70
EX SP
CsgA
BBa_K1404006
BIO ENGINEERING
CsgA- strain
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CsgA p70
EX SP
CsgA
BBa_K1404006
BIO ENGINEERING
CsgA- strain
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CsgA p70
EX SP
CsgA
BBa_K1404006
BIO ENGINEERING
CsgA- strain
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BBa_K1404006
OmpR234 strain
BIO ENGINEERING
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CsgA p70
EX SP
BBa_K1404006
OmpR234 strain
BIO ENGINEERING
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CsgA p70
EX SP
BBa_K1404006
BIO ENGINEERING
OmpR234 strain
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BBa_K1404007
BIO ENGINEERING
OmpR234 strain
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CsgA
p70
EX SP
His 1
BBa_K1404007
BIO ENGINEERING
OmpR234 strain
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CsgA
p70
EX SP
His 1
BBa_K1404007
CsgA His 1
BIO ENGINEERING
OmpR234 strain
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CsgA
p70
EX SP
His 1
BBa_K1404007
CsgA His 1
BIO ENGINEERING
OmpR234 strain
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CsgA
p70
EX SP
His 1
BBa_K1404007
CsgA His 1
BIO ENGINEERING
OmpR234 strain
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CsgA: THE MAIN CURLI SUBUNIT
WILD TYPE CsgA
Without any Tag
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CsgA His1
CsgA: THE MAIN CURLI SUBUNIT
WILD TYPE CsgA
Without any Tag
His 1
With one His Tag
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Without any Tag
WILD TYPE CsgA
CsgA: THE MAIN CURLI SUBUNIT
Nickel
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Without any Tag
WILD TYPE CsgA
+ 25 %
With one His Tag
CsgA His1
CsgA: THE MAIN CURLI SUBUNIT
Nickel
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Nickel
CsgA: THE MAIN CURLI SUBUNIT
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CsgA x1 Nickel
CsgA: THE MAIN CURLI SUBUNIT
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CsgA x1 Curli x1 Nickel
CsgA: THE MAIN CURLI SUBUNIT
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CsgA x1 Curli x1 CsgA x n Nickel
CsgA: THE MAIN CURLI SUBUNIT
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Bacteria CsgA x1 Curli x1 CsgA x n Nickel
CsgA: THE MAIN CURLI SUBUNIT
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Curli x n Bacteria CsgA x1 Curli x1 CsgA x n Nickel
CsgA: THE MAIN CURLI SUBUNIT
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Curli x n Bacteria CsgA x1 Curli x1 CsgA x n Nickel
CsgA: THE MAIN CURLI SUBUNIT
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Curli x n Bacteria CsgA x1 Curli x1 CsgA x n Nickel
CsgA: THE MAIN CURLI SUBUNIT
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His CsgA
HisTag: TWO POSSIBLE CONFORMATIONS
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His CsgA His CsgA
HisTag: TWO POSSIBLE CONFORMATIONS
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His 1 Polymerized curli
MODELING HYPOTHESIS
Curli polymerization
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His 1 Polymerized curli His 1 CsgA
Ni Ni Ni Ni
MODELING HYPOTHESIS
Curli polymerization With His Tag
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His 1 Polymerized curli His 1 CsgA
Ni Ni Ni Ni
until His n … His 1 His 2 His 3
Ni Ni Ni Ni Ni Ni Ni Ni Ni Ni Ni Ni
CsgA
MODELING HYPOTHESIS
Curli polymerization With His Tag
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No TAG
RBS
CsgA
CsgA
OUR BIOBRICKSTM
BBa_K1404006
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No TAG
RBS
CsgA
CsgA His 1
RBS
CsgA His1
CsgA
OUR BIOBRICKSTM
BBa_K1404006 BBa_K1404007
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No TAG
RBS
CsgA
CsgA His 1
RBS
CsgA His1
CsgA
RBS
CsgA
His1 His2
His 1 His 2 CsgA
OUR BIOBRICKSTM
BBa_K1404006 BBa_K1404007 BBa_K1404008
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NEGATIVE CONTROL NO CURLI
CURLI STRUCTURE
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NEGATIVE CONTROL NO CURLI POSITIVE CONTROL E.coli + BBa_K1404006 (No His Tag)
CURLI STRUCTURE
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NEGATIVE CONTROL NO CURLI POSITIVE CONTROL E.coli + BBa_K1404006 (No His Tag) CURLI FORMATION
CURLI STRUCTURE
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E.coli + BBa_K1404007 (1 His Tag)
CURLI STRUCTURE
POSITIVE CONTROL E.coli + BBa_K1404006 (No His Tag)
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E.coli + BBa_K1404007 (1 His Tag) STRONG CURLI FORMATION
CURLI STRUCTURE
POSITIVE CONTROL E.coli + BBa_K1404006 (No His Tag)
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E.coli + BBa_K1404007 (1 His Tag) E.coli + BBa_K1404008 (2 His Tags) STRONG CURLI FORMATION
CURLI STRUCTURE
POSITIVE CONTROL E.coli + BBa_K1404006 (No His Tag)
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E.coli + BBa_K1404007 (1 His Tag) E.coli + BBa_K1404008 (2 His Tags) STRONG CURLI FORMATION CURLI FORMATION
CURLI STRUCTURE
POSITIVE CONTROL E.coli + BBa_K1404006 (No His Tag)
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NEGATIVE CONTROL
(CsgA- strain)
BIOFILM STRUCTURE
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NEGATIVE CONTROL
(CsgA- strain)
E.coli + BBa_K1404006 (No His Tag)
BIOFILM STRUCTURE
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E.coli + BBa_K1404007 (1 His Tag) E.coli + BBa_K1404006 (No His Tag) E.coli + BBa_K1404008 (2 His Tags)
BIOFILM STRUCTURE
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E.coli + BBa_K1404007 (1 His Tag) E.coli + BBa_K1404006 (No His Tag) E.coli + BBa_K1404008 (2 His Tags)
BIOFILM STRUCTURE
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10 20 30 40 50 60 70 80 90 100 CsgA- (negative control) No Tag (positive control) His 1 His 2
Percentage of Adherence
Adherence (%)
Crystal Violet Coloration
ADHERENCE
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10 20 30 40 50 60 70 80 90 100 CsgA- (negative control) No Tag (positive control) His 1 His 2
Percentage of Adherence
Adherence (%)
Crystal Violet Coloration
ADHERENCE
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Ni Ni Ni Ni Ni Ni Ni Ni Ni Ni
BIOFILM MATRIX
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48 hours E.coli biofilm E.coli liquid culture
NICKEL CHELATION: PROCEDURE
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48 hours E.coli biofilm Add Ni2+ E.coli liquid culture
NICKEL CHELATION: PROCEDURE
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48 hours E.coli biofilm Add Ni2+ E.coli biofilm containing Ni2+ Nickel chelation : 30 min E.coli liquid culture
NICKEL CHELATION: PROCEDURE
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48 hours E.coli biofilm Add Ni2+ E.coli biofilm containing Ni2+ Nickel chelation : 30 min Ni Concentration: ICP – MS E.coli liquid culture
NICKEL CHELATION: PROCEDURE
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0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 WT HIS 1 HIS 2
Determination of Nickel chelated
µgNi / AU Curli
NICKEL CHELATION: RESULTS
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0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 WT HIS 1 HIS 2
Determination of Nickel chelated
µgNi / AU Curli
NICKEL CHELATION: RESULTS
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0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 WT HIS 1 HIS 2
Determination of Nickel chelated
µgNi / AU Curli
NICKEL CHELATION: RESULTS
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His 1 CsgA
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His 1 CsgA
Ni Ni Ni Ni Ni Ni Ni Ni Ni Ni
CsgA His 2
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OUR HP INSPIRED SOLUTION: FILTER CURLY’ON
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Polluted water
OUR HP INSPIRED SOLUTION: FILTER CURLY’ON
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Polluted water Purified water
OUR HP INSPIRED SOLUTION: FILTER CURLY’ON
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Inside of the filter Polluted water Purified water
OUR HP INSPIRED SOLUTION: FILTER CURLY’ON
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Inside of the filter Polluted water Purified water
OUR HP INSPIRED SOLUTION: FILTER CURLY’ON
Damien Jacques, Design and Mechanical Engineering Baptiste Laubie University of Lorraine Expert in Recycling EXPERTS
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HUMAN PRACTICE: sociological inquiry
Recycling Food Processing Dialysis Inox industry water depollution
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10 20 30 40 50 60 70
WHICH PURIFICATION TECHNIC(S) DO YOU PREFER FOR DRINKING WATER ?
HUMAN PRACTICE: SURVEY
Biological method (proteins) Alive bacteria Alive GM bacteria Physical method Chemical Method
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10 20 30 40 50 60 70
WHICH PURIFICATION TECHNIC(S) DO YOU PREFER FOR DRINKING WATER ?
HUMAN PRACTICE: SURVEY
Biological method (proteins) Alive bacteria Alive GM bacteria Physical method Chemical Method
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HUMAN PRACTICE: SURVEY
10 20 30 40 50 60 70 PEOPLE SURVEYED (%)
Have you heard about synthetic biology?
YES NO
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10 20 30 40 50 60 70
WHICH PURIFICATION TECHNIC(S) DO YOU PREFER FOR DRINKING WATER ?
HUMAN PRACTICE: SURVEY
Biological method (proteins) Alive bacteria Alive GM bacteria Physical method Chemical Method
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10 20 30 40 50 60 70
WHICH PURIFICATION TECHNIC(S) DO YOU PREFER FOR DRINKING WATER ?
HUMAN PRACTICE: SURVEY
Biological method (proteins) Alive bacteria Alive GM bacteria Physical method Chemical Method
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No UV exposure
After 10 min UV exposure After 15 min UV exposure After 20 min UV exposure
SAFETY: UV treatment
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Healthy bacteria Altered bacteria Dead bacteria No UV exposure
After 10 min UV exposure After 15 min UV exposure After 20 min UV exposure
SAFETY: UV treatment
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UV Ethanol Heating Degradation of genomic DNA Death of bacteria
SAFETY: our complete solution
Death of bacteria
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UV Ethanol Heating Degradation of genomic DNA Death of bacteria
SAFETY: our complete solution
Death of bacteria
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n
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n
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n
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n
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n
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n Nanobodies
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MAP and BF2I Laboratories Biosciences Department
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