Biophysics – Biological Physics
● nomenclature
● fields of research
● history: old discipline, turning point is recent
● why physicists in biology?
● new multidisciplinary field: system biology, synthetic biology
● biophysics at in the Physics department at UMN
● in vitro gene expression and protocell
● biological physics: education, courses, resources.
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Biophysics / Biological Physics
Biophysics:
- more used among physiologists, biochemists
- molecular level
Biological physics:
- preferred by physicists
- molecular to ecological
Ideas of biophysics or biological physics are the same:
- fundamental physics of biological systems and processes.
- apply the techniques from physics to understand biological
structure and function.
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Biophysics, major components / examples
- explain biological function in terms of molecular mechanisms
- ions channels
- protein 3D structures and functions (crystallography)
- DNA replication
- conversion of external signals to electrical signal
- conversion of chemical energy to mechanical force (muscle)
ATP hydrolysis = 20 kBT
1 kBT =1 pN nm
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Biological physics, major components / examples
- molecular: channels in membranes, dynamics of chemical reactions
- subcellular: transport, signal processing, dynamics of
polymerization, motility, flagellar dynamics
- cellular: chemotaxis, swimming, crawling, growth
- multicellular: pattern formation, morphogenesis
- organism: cardiac dynamics, circadian rhythms, information
processing
- evolution / ecology: in vitro evolution, population dynamics
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History
Ideas are not new:
● D’Arcy Thompson (1860-1948) talking about cells, tissues,
bones, flowers: “Their problems of form are in the first instance
mathematical problems, their problems of growth are essentially
physical problems.”
● E. Schroedinger: “what is life?” (1944).
Recent field in terms of research effort:
- became clear in the past decade: explosion of meetings, journals.
- many departments building research groups in biophysics /
biological physics.
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Why physicists in biology?
What physicists can bring:
- quantitative measurements
- modeling and testing
- reductive approaches
- universality of behavior
- development of new methods and technologies
- in vitro / synthetic approaches
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Biophysics Group
Theory:
Alexander Kamenev
Boris Shklovskii
Experiment:
John Broadhurst
David Thomas
Joachim Mueller
Vincent Noireaux
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Prey Population
Alex Kamenev
1. Populations dynamics,
as an example of non-equilibrium
statistical mechanics.
Predator Population
2. Transport through ion channels,
as an example of 1D physics.
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Electrostatic theory of viral self-assembly
Boris Shklovskii
r
R
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Thomas Lab
Spectroscopic Probes of Muscle Protein Structure and Dynamics
Myosin
Cardiac
Calcium
Pump
ATP
ADP + Pi
Ca2+
ATP
ADP + Pi
Phospholamban
Ca2+
Actin
Probes
Spectroscopic Probe Methods:
Electron paramagnetic resonance (EPR)
Nuclear Magnetic Resonance (NMR)
Time-resolved fluorescence and phosphorescence
O
O
H
N C CH2I
•N
I
HO
I
O
O
I
I
COOH
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N=C=S
J. Broadhurst
Magneto encephalography (MEG)
Study the location in the human brain of the processors of external stimuli.
Currently work is being done on the identification of
different sounds by a part of the brain above and in front of the
ear. (This part is known as the auditory cortex, and is located in a
fold of the brain called the sylvan fissure).
When a sound is received by the ear, it is analyzed into the
different frequencies that it contains, before being passed on to
the first level of processing. This identifies loudness and the
direction of the sound source, and then transmits the information
to the second processor, which tries to identify the identity of the
sound (Is it a violin, or a cat meowing?)
Neurons in the brain
activate and produce tiny
magnetic fields (10-12 Tesla ).
An array of 250 superconducting magnetometers
(squids) are used to measure
the fields.
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Mueller Lab: Fluorescence Fluctuation Spectroscopy (FFS)
Two-Photon
Spot
objective
Single-molecule microscope
FFS in cells
Two-photon Effect
Watch Protein Interactions in Living Cells:
Concentration (nM)
2
10000
4
10
10
dimer
9000
8000
app(cpsm)
Photon
Count
Statistics
3
10
7000
RAR LBD
TR4
6000
5000
4000
3000
12
monomer
2000
4
10
10
5
Intensity (Counts per second)
10
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Joachim Mueller:
Viruses
Protein Assemblies and
Fluctuation
Analysis
2-photon
spectroscopy
Light burst from single
molecules passing through
tiny optical volume
Image of a cell assembling viral-like
particles. We study assembly process of
retroviruses, such as HIV-1.
Construct physical model
of assembly pathway
Harvest viral
particles
Fluctuation
Analysis
5500
Intensity (cps)
5000
4500
Viral
Particle
4000
3500
3000
2500
2000
0
100
200
time (sec)
Microfluidics of viral particles
300
400
• protein coat contains holes
• the hole density varies
• below percolation threshold
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Vincent Noireaux
● information processes (synthetic genetic circuits).
● biopolymer self-assembly at the membrane: cell division, motility, nano by bio.
● artificial cell system.
Reconstitution of genetic circuits in vitro.
Coarse-grained model of circuits.
Artificial cell system.
Self-assembly of proteins/biopolymers.
20μm
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Vincent Noireaux
● information processes (synthetic genetic circuits).
● biopolymer self-assembly at the membrane: cell division, motility, nano by bio.
● artificial cell system.
genome DNA of virus
cell-free expression in test tube
de novo synthesis of virus
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Biophysics courses at UMN
● Physics department:
- 4911/5081: intro. to biopolymer physics.
- 5401: physiological physics.
- 5402: radiological physics.
● other courses:
- Math 5445: mathematical analysis of biological networks.
- Math 8540: topics in mathematical biology.
- biology courses.
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DNA sequencing and synthesis
Sequencing of bacterium genome: 1 week (5 Mb)
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Information
 man-made
nature - evolution 
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New interdisciplinary fields
● system biology:
(1) understanding the structure of the system, such as gene regulatory
networks.
(2) understanding the dynamics of the system, both quantitative and
qualitative analysis.
(3) understanding the control methods of the system.
(4) understanding the design methods of the system, are key milestones to
judge how much we understand the system.
● synthetic biology: the design and fabrication of biological components and
systems that do not exist in the natural world. Use them either as molecularscale factories, to make simple computations, deliver vaccines, or to create
new hybrid materials. Like system biology, synthetic biology is at a very
preliminary stage but physicist could have a significant scientific impact.
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Molecular programming in a
test tube: synthetic gene
circuits, phage synthesis and
artificial cell.
Vincent Noireaux, UMN
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Introduction – Motivations
• The three components of cellular life.
• the bottom-up approach to living systems.
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Living cell
(bacteria E. coli)
DNA  RNA  proteins
Genome (DNA):
- 5 millions bases
- 4500 genes
- hundreds gene circuits
Genome (DNA)
Nutrients
1 μm
(E. coli)
Self-reproduce in 30 min.
Capable of:
- responding to stresses
- sensing the environment
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Cell: the basic unit of life
Information
Compartment
Metabolism
Unique property: self-reproduction.
Each part is essential.
Each part is made of molecular machineries.
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Synthetic biology era
The design and fabrication of biological components and systems
that do not exist in the natural world:
• to understand gene regulation and make simple computations.
• to use them either as molecular-scale factories.
• to create new hybrid materials.
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Synthetic biology platforms
in vivo
in silico
in vitro
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Synthetic biology in a test tube
(cell-free synthetic biology)
Constructing living systems in a test tube
from the DNA program.
DNA
TX
mRNA
TL
protein
● bottom-up, reductionist and constructive approach.
● no endogenous information.
● no interference and response from an organism.
● more freedom of control and design compared to in
vivo.
● molecular programming approach to living systems.
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2
Small gene circuits in a test tube
(DNA
TX
mRNA
TL
protein)n
circuits
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Transcriptional activation cascade
σ70
P70
σ28
P28
deGFP
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AND gate S54-NtrC
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Multiple stage cascade
σ70
P70
σ38
P38
σ19
P19
σ28
P28
T7rnap
PT7
deGFP
m  12  2 min
Leak!
Loss of specificity

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Multiple stage cascade
σ70
P70
σ38
P38
σ19
P19
σ28
P28
T7rnap
PT7
deGFP
m  6  1min

Leak attenuation
Specificity
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Conclusion:
• constructed and characterized cell-free circuits.
• learned the design rules.
• tuned the dynamics.
• global mRNA degradation rate is critical.
• Shin and Noireaux. ACS Synthetic Biology 2011.
Information
Compartment
Metabolism
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Genome scale circuits
(information and self-organization)
● What is the real capacity of the system to construct
circuits
and living systems?
50-60 genes
CFR batch mode: [Protein] = 25-30µM
E. coli: [Protein]ave = 500nM
● Test the system with genome-sized information.
● Bacteriophages:
- search for genomes composed of ≤ 60 genes.
- with molecular biology technically accessible.
- condition/bottleneck: complexity of the interaction with the
host beyond TX-TL.
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Phage T7
●
●
●
●
●
lytic coliphage.
40 kbp, 60 genes (35 with known functions).
almost host independent (2 host proteins required).
has its own RNA polymerase.
has its own DNA polymerase.
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Phage T7 synthesis in a test tube
TX
genome
TL
mRNA
phage
● TEM image
● 5-6 hours of
incubation
● batch mode reaction
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T7 Genome replication
● up to 200 times greater
with dNTPs.
● a
few
billion
of
functional phages per
milliliter synthesized after
5-6 hours of incubation in
batch mode.
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T7 - E. coli Infection test
No difference observed between in vivo and in vitro
synthesized phages.
● phages per cell ≈ 100.
● phage cycle ≈ 25 min.
● E. coli division ≈ 30
min.
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