***This research has been published in Science, Geochimica et Cosmochimica Acta, Advances in Agronomy, Geomicrobiology Journal, and Eos.
bacterium-mineral interfaceinterfacial forces and proteins
*forces control, and are themselves modulated by, the expression of biopolymers on a bacterium’s surface
400 nm
G. B
owles
A comparison of two organisms that work in the realm of the nanometer.
1030 109
106 100
109 106
Microorganisms Humans# of cells
# of species
# years
Consider the
following…
Shewanella interactions with Fe oxyhydroxidesmetal reducing bacteria & oxides
primitive form of respiration using Fe(III) solids
remediation of organic and inorganic pollutants in surface and subsurface environments
Shewanella
Fe(III) mineral
2D gel electrophoresis
pHMr
phage display library
force microscopy
cell lithography
discover forces and proteins
mimic and utilize information
Cell-material interface
F~e -D
theory
-1-0.8-0.6-0.4-0.2
00.2
0 100 200 300 400 500 600
distance (nm)
forc
e (n
N)
Force-distance curves using different minerals, bacteria, and solutions
approach
retraction
G x m1 x m2
(distance)2
electrostatic and van der Waals forcesDLVO Theory
σ = surface charge R = radius ε = dielectric constant εo= vacuum permittivity d = distance κ = 1 / Debye length
F(d) =4 π σ1 σ2 R
ε εo κ e - κ d H R6 d 2
H = Hamaker constant R = radius d = distance
~(salt concentration) –1/2
green – low IS
blue – high IS
subsurface transport in the environment(approach measurements between G- bacterium and a silicate)
-1.000.001.002.003.004.005.00
0.0 20.0 40.0 60.0 80.0 100.0distance (nm)
forc
e (n
N/ µ
m)
approach only
-1-0.8-0.6-0.4-0.2
00.2
0 100 200 300 400 500 600distance (nm)
forc
e (n
N)
goethitediaspore
Force-distance curves (retraction forces between Shewanella and AlOOH vs FeOOH)
approach
retraction
AlOOH FeOOH
Shew
energy values between Shewanella and mineral (as function of oxygen concentration)
attoJoules (10-18 J)
anaerobic
aerobic
diaspore(AlOOH)
137 20+-
26 6+-
goethite(FeOOH)
41 4+-
+-39 7
Control experiment with nonviable cells ~6 aJ (did not change with mineral, oxygen concentration, or contact time)
Protein folding/unfoldingWorm-like Chain Model
F(d) = (k T / b) [0.25 (1 – d / L)–2 – 0.25 + d / L]
d = distance or extension k = Boltzmann’s constant T = temperature b = persistence length (0.38nm Cα-Cα in protein) L = contour length (length of stretched protein chain)
-1-0.8-0.6-0.4-0.2
00.2
0 100 200 300 400 500 600distance (nm)
forc
e (n
N)
150 kDa
goethitediaspore
OM protein expression patternsShewanella and AlOOH or FeOOH
protein signature observed in 80% of the data; only with goethite after some period of “recognition time”
excise proteins& fragment
into peptides
pHMr
pH
Mr
mass spec & database search
2D Gel Electrophoresis of OM Proteins
extract membrane
proteins
culture cells
separate proteins
compare with force signature
-1-0.8-0.6-0.4-0.2
00.2
0 100 200 300 400 500 600
distance (nm)
forc
e (n
N)
150 kDa
anaerobicaerobic
force microscopy
Fe+3
O2
2D gel electrophoresis
pHMr
phage display library
force microscopy
cell lithography
discover forces and proteins
mimic and utilize information
Cell-material interface
F~e -D
theory
peptide phage library expose to target mineral
wash away unbound phage elute bound phage “bio-panning”
isolate clones, sequence DNA to find mineral-binding motif,
biological cell lithographybiological cell lithography
substrate
cell
protein
bacterium as living “pen” that produces and secretes genetically engineered proteins
T. Beveridge
S-layer protein
bacterium-mineral interfacenanoscale forces and proteins
• quantify natural forces of affinity between inorganic crystalline phases and proteins synthesized by bacteria
• use theoretical models and protein expression patterns to identify putative mineral specific proteins
• mimic natural specificity by attempting to fabricate peptides with unique mineral-binding motifs
• use living microbial cells as a lithographic tool
AcknowledgementsAcknowledgementsTerry Beveridge, John Smit, Courtney Crummett, Graeme Bowles
Steven Lower – Ohio State University – [email protected]
National Science Foundation (GEO & ENG)
Department of Energy
American Chemical Society***This research has been published in Science, Geochimica et Cosmochimica Acta, Advances in Agronomy, Geomicrobiology Journal, and Eos.