Nanobiotechnology
Petr Skládal
Department of Biochemistry, MU
RG & CF Nanobiotechnology, CEITEC
youtube.com
Nanobiotechnology at MU
at MU, we have started research in this area around
2005 (purchase of our first AFM Ntegra Vita at NCBR)
significant expansion within CEITEC – Structural
Biology research program – the Core Facility of
Nanobiotechnology Tools
… we need to provide introduction to the future users
of techniques available at this CF, as well as to
educate our (PhD) students
course is also incorporated to the Life Science PhD
program at CEITEC, but opened to all students from
MU
Lecturers: Petr Skládal, Jan Přibyl, Antonín Hlaváček,
Karel Lacina
Sylabus
1. Introduction. Science of Nano. What is nanobiotechnology.
2. Nanostructures. Carbon nanotubes, semiconductor nanoparticles quantum
dots. Metal-based nanostructures - nanowires and bioelectronics.
Gold nanoparticles (nanorods, nanocages, nanoshells). Magnetic
nanoparticles. Polymer nanostructures (dendrimers). Protein-based
nanostructures - nanomotors from microbes and mammalian cells
(myosin). Nanomachines based on nucleic acids.
3. Experimental technichues. Scanning probe microscopies (STM, AFM,
SNOM, SECM, ...). Physical principles, basic and advanced measuring
modes. Imaging of bioobjects - from atoms, molecules to cells and tissues.
Combined techniques with inverted optical and fluorescence microscopes.
Raman imaging. Biointeractions at the molecular level.
4. Self-assembling techniques. Separation, characterization and
modification of nanoparticles. From natural to artificial structures.
Nanolithography and nanomanipulations. Nanoparticles for biological
labeling and cellular imaging. Nanobiosensors and nanobioanalytical
systems. Microfluidics, cell sorting and lab-on-a-chip. Biochips and
sensing arrays, nanodeposition of biomolecules.
5. Medical applications. Cytotoxicity of nanoparticles. Nanostructures in
drug discovery, delivery and controlled release. Nanostructures in cancer
research. Nanotechnology for tissue engineering and regenerative therapy.
6. Nanobiotechnology in commercial examples. Perspectives and
conclusions.
… might be modified …
Brief History
“There is plenty of room at the bottom.”
R. Feynman, 1959 Caltech; Father of Nanotechnology
Dr. Richard P. Feynman (1918-1988)
nano, Greek for “dwarf,” means
one billionth
The principles of physics, as far
as I can see, do not speak
against the possibility of
maneuvering things atom by
atom. It is not an attempt to
violate any laws; it is something,
in principle, that can be done;
but in practice, it has not been
done because we are too big”
There’s Plenty of Room at the Bottom: An Invitation to
Enter a New Field of Physics – by Richard P. Feynman
the term “nanotechnology” appeared few years after Feynman’s lecture
his remarks are still relevant half a century after his original address
related to us is Feynman’ vision regarding the field of biology. Unlike
many physicists of his time, Feynman realized how important biology is in
the framework of nanotechnology
The marvelous biological system.
“The biological example of writing information on
a small scale has inspired me to think of something
that should be possible. Biology is not simply writing
information; it is doing something about it.”
Rearranging the atoms
“But I am not afraid to consider the final question as
to whether, ultimately---in the great future---we can
arrange the atoms the way we want; the very atoms,
all the way down! What would happen if we could
arrange the atoms one by one the way we want them”
Engineering & Science magazine, California Institute of Technology, Vol. 23/5, Feb. 1960
Richard Smalley
a chemistry professor at Rice University, pioneered
the field of nanotechnology and shared a Nobel Prize
in 1996 for the development of bucky-balls
(fullerene)
Dr. Richard E. Smalley (1943-2005, 1996 Nobel Prize in Chemistry)
Fullerenes
a new form of carbon clusters was identified in 1985, graphite
was vaporized using a laser
closed and convex cage molecules formed by the arrangement of
hexagonal and pentagonal faces
buckminsterfullerene, named after the architect R. Buckminster
Fuller, C60, size is about 0.7 nm, excellent electrical and heat
conductivity
fullerenes that have 70, 76, 84, 90, and 96 carbon atoms were also
found
smallest possible fullerene, C20, consists of solely of 12
pentagons, generated from a brominated hydrocarbon,
dodecahedrane, by gas-phase debromination, lifetime 0.4 ms
potential use as nanocontainers, some antiviral activity
elongated variants – carbon nanotubes (1991, Sumio Iijima),
single and multiwall forms (SWCNT and MWCNT)
Eric Drexler
as a researcher, author, and policy advocate he has been one of
the pioneers to focus on emerging technologies and their impact
for the future
founded the Foresight Institute, presently at Nanorex, a
company that develops software for the design and simulation
of molecular machine systems
his thought provoking publications
– “Engines of Creation: The Coming Era of Nanotechnology,”
– “Nanosystems: Molecular Machinery, Manufacturing,
and Computation,”
– “Unbounding the Future:
The Nanotechnology Revolution”
made great impact by introducing the very
topic of nanotechnology to many
Dr. Eric Drexler (1955-)
Two-week revolution
Drexler proposed constructing molecules by forcibly pressing
atoms together into the desired molecular shapes “mechanosynthesis”
(parallels with macroscopic machinery
and engineering)
building objects in an assembly-line manner by directly bonding
individual atoms
central idea - construction of an assembler, a nanometer-scale
machine that assembles objects atom-by-atom according to
defined instructions
creation of just a single working assembler would lead immediately
to the “Two-Week Revolution
still only an idea …
other way - nanotechnology that looks to nature for its start; cells
build thousands of working nanomachines, which may be
harnessed and modified to perform our own custom
nanotechnological tasks = Nanobiotechnology
a slower process, but feasible
Tools for nanobio
bottom-up vs. top-down approach
building innovative tools to study and manipulate
biology at the nanometer scale
– new tools
– new materials
– new devices
– new knowledge
http://www.nanotech-now.com/columns/?article=097
Scanning probe microscopies
invention of the scanning tunneling microscope
(STM) by Gerd Binnig and Heinrich Rohrer at IBM’s
Zurich Research Labs (1986 Nobel Prize in Physics)
Followed five years later by the invention of the
atomic force microscope (AFM)
Gerd Binning Heinrich Rohrer
tip
applied V -> I
I ~ V exp(-AΦΦΦΦ1/2d)
Chad Mirkin
Professor in the Institute for Nanotechnology at Northwestern
University
a pioneer in chemical modifications of nanosystems leading to
breakthrough contributions to bionanotechnology
explains the need to open our minds and change our attitude as
we embark on learning this new field:
“At the nano level atoms do not belong to any
field of science. ”
this conveys the extreme diversity and
uniqueness of nanotechnology, while
stressing the preparation required by
those aspiring to contribute to it
developed dip-pen nanolithography
technique, uses AFM to introduce
patterned molecules into surface
Dr. Chad Mirkin (1963-)
Definition of nanotechnology
Nanotechnology can be difficult to determine and define
realm of nanoscience is not new; chemists will tell you
they’ve been doing nanoscience for hundreds of years
stained-glass windows found in medieval churches
contain different-size gold nanoparticles incorporated
into the glass — creating orange, purple, red, or
greenish colors.
Einstein, as part of his doctoral dissertation, calculated
the size of a sugar molecule as one nanometer
loosely considered, both the medieval glass workers
and Einstein were nanoscientists …
Nanotechnology
research and technology development at the atomic, molecular or
macromolecular levels, in the length scale of approximately 1 - 100
nanometer range
creating and using structures, devices and systems that have
novel properties and functions because of their small and/or
intermediate size.
ability to control or manipulate on the atomic scale
… National Nanotechnology Initiative, USA
… alternatively:
design, characterization, production and application of structures,
devices and systems by controlling shape and size at nanometer
scales
Nanoscience
study of phenomena and manipulation of materials at atomic,
molecular and macromolecular scales, where properties differ
significantly from those at a larger scale
Nanobiotechnology
applies tools and processes of nano / microfabrication to build
devices for studying biosystems
biotechnology is the application of technological innovation as it
pertains to biological and life sciences. Nanobiotechnology
incorporates biotechnology on the nano-scale
is a subset of nanotechnology where the biological world
provides
the inspiration and/or the end goal. It is defined as atom-level
engineering and manufacturing using biological precedence for
guidance (Nano-Biomimetics) or traditional nanotechnology
applied to biological and biomedical needs
slightly different is Bionanotechnology
use of biological building blocks and the utilization of biological
specificity and activity for the development of modern technology
at the nano-scale
focus on biological nanomachines
Wikipedia:
Bionanotechnology generally refers to the study of how the goals
of nanotechnology can be guided by studying how biological
"machines" work and adapting these biological motifs into
improving existing nanotechnologies or creating new ones
Nanobiotechnology, on the other hand, refers to the ways that
nanotechnology is used to create devices to study biological
systems
nanobiotechnology is essentially miniaturized biotechnology,
whereas bionanotechnology is a specific application of
nanotechnology
for example, DNA nanotechnology or cellular engineering would
be classified as bionanotechnology because they involve working
with biomolecules on the nanoscale
conversely, many new medical technologies involving
nanoparticles as delivery systems or as sensors would be
examples of nanobiotechnology since they involve using
nanotechnology to advance the goals of biology
Nanobio- or Bionano- technology
Bionanotechnology as the use of biological assemblies for various
applications that may not be directionally associated with biology.
Nanobiotechnology is the use of nano-science for specific biological
applications
Nanobiology
this can serve as a description of both bionanotechnology and
nanobiotechnology
Bionics, which stems from [bi(o)- + (electr)onics] could be
described as the application of biological principles and
mechanisms to the design and fabrication of engineering
systems
not merely to copy nature but rather to understand its principles
and use them as a stimulus and motivation for innovations
bio-inspired technology may be more suitable name
Nanobionics is applied at the nanoscale level
Convergence of disciplines
and technology transfer from physics to the bio/medical sciences
to solve Grand Challenges
molecular:
– structure analysis of single proteins (attosecond
spectroscopy)
– sequencing single DNA (nanopores)
– understanding DNA damage and repair
cellular:
– molecular scale imaging of single, living cells
– single molecule biochemistry in single, living cells
medical:
– detecting single, abnormal cells among healthy ones in living
tissues
– developing non-invasive medical tools
– understanding the cellular biochemistry of the brain
How small is small?
From macro to nano…
macro (m) meso (mm) micro (um) nano (nm)
bridges
airplanes
cars
human beings
birds
snails
rice
ants
watch gears
sand
hair
dust
pollen
cells
bacteria
viruses
nuclei
transistors
cell apparatus
proteins
Object Diameter (nm)
Hydrogen atom 0.1
Water molecule 0.3
DNA (width) 2.5
Cell membrane 5-9
Porin channel 4-10
Actin filament 5-9
Microtubule 25
Bacterial flagellum 12-25
Magnetosomes 35-150
Liposome vesicle 100 (min)
Bacterial cell 250 - 1,000
Spores 1,000 - 8,000
Red blood cell 6,000 - 8,000
Human hair 100,000
Cockroach Human
Hair
Pollen
Grain
Red
Blood
Cells
Cobalt
Crystal
Lattice
Palladium
Half-
shells
Aspirin
Molecule
Nanobiotech
systems
some examples
from different
fields
Scaling effects
If a system is reduced isomorphically,
the changes in length, area and volume
ratios alter the influence of various
physical effects that determine the
overall operation
Gravity and inertia are
negligible at the nanoscale
macroscopic objects - dominated by properties of mass
– cm to m sizes - physical properties such as friction, tensile strength,
adhesion, and shear strength are comparable to inertial and gravity forces
– this balance changes, however, for larger or smaller objects
– increase in inertia or weight can quickly overcome the increase in strength in
a large structure such as a building (common sense to add extra support)
opposite effect for smaller and smaller objects
– um-sized objects (nanoparticles, individual cells) interact differently
– inertia is no longer a relevant property
actions of small objects are dominated by their interaction with
neighboring objects
– fine dust stays suspended in the air instead of dropping quickly
– nano-objects in water undergo random Brownian motion
– attractive forces between small objects are stronger than gravity
Scaling laws
Typical small values
Some small scale
phenomena
At the atomic level, we have new kinds of forces and
new kinds of possibilities, new kinds of effects.
Richard Feynman
Surface tension
Laminar flow
Rapid heat transfer
Surface area / volume
for a small molecule D≈≈≈≈5x10-5 m2/s
mixing by diffusion is very fast at the micro-level
In nature animals made up of more the a few cells cannot rely on diffusion
any more to move materials within themselves. They augment transport with
hearts, blood vessel, etc.
Scaling and diffusion
Oxygen storage
macro level
– high pressure tanks
– delivered in a continuous
stream in tubes
– flow controlled by valves
nano level
– transported by individual molecules, low pressure conditions
– molecules meet with the carrier – hemoglobin by random diffusion,
resulting in formation of a complex
– co-operative effects allow to gather oxygen efficiently when its
content rise and discharge oxygen completely when its level drops
Loss of continuity
Phenomena at the nanometer scale:
atomic granularity
Surface forces
Fast time scales
Some VERY small scale phenomena:
Noise
How to deal with small objects:
techniques to reduce adhesion
Engineering small: How small is
small enough?
Engineering small: How small is
small enough?
sub-miniature (~ 1 mm)
conventional precision engineering, primarily metals, glass
micro-electrical mechanical systems (MEMS~10-100 µm)
sensors and “useful” devices, primarily in silicon
nano-electrical mechanical systems (NEMS~100-1000 nm)
electronic devices, concept devices, primarily in silicon
nanotechnology (<100 nm)
mostly materials
different perspectives, different communities
Engineered systems
materials
basic building stuff, often with special properties
processed in batch
components
simple parts designed to be put together
devices
assembly of components that can perform a simple
function
sub-systems (modules)
assembly of devices designed to perform a complex
function
systems
assembly of sub-systems designed to perform desired
application
how is a system engineered today?
Engineered systems
Example system: automobile
Final system:
automobile
Sub-systems
Device (brake)
Component
Material
Engineered systems
Example system: human body
System: Human body
Device
(Ribosomes) Material
(amino acids)
Sub-systems
(organs, cells)
Component
(proteins)
Engineered systems with nanotech
Engineered systems can take us to wonderful places…
Enabled by very small technology
The challenges of
nanobiotechnology
Spatial fidelity
molecule < molecular structures < organelles < cells
1 nm 10 nm 100 nm 1 um 10 um
proteins coated pits - caveolae mitochondria nucleus
Imaging:
atomic force microscopy
electron microscopy
optical microscopy
Spectroscopy:
optical (fluorescence) spectroscopies
mass spectrometry
Time scale of events
ps - bond vibrations
triggering physics
ns - conformations
us - binding associations
chemistry
ms - reactions flux
s - regulation transport
ks - movement regeneration
biology
Ms - development fetal Growth
Gs - life cycle disease
Heterogeneity
many compartments, many structures, many surfaces …
as many as 85% of biochemical interactions occur at or in a membrane
Complexity
a small number of needles:
1,000 – 100,000 copies of a particular protein
a few percent are modified at any given time - a few
copies per cell?
in a very large hay stack:
~ 30,000 genes
~ 100,000 gene products
> 1,000,000 post-translationally modified proteins
must study specific intermolecular interactions in a mixture
Specificity
measure pair-wise interactions systematically - sequentially
- exclusively and whether transient or long lived
A-A interactions
A-B interactions
Sensitivity typical
Concentrations in a cell or Densities on a cell surface
N = 100,000 molecules
V = 10x10x10 um3
= 1000 fL = 1 pL
c = 100 molecules per um3
c ~ 100 nM
N = 100,000 molecules
A = 10x10x6 um2
= 600 um2
c ~ 100 molecules per um2
we need single molecule detection sensitivity
to achieve sub-micron resolution
fluorescence can provide 100 counts per molecule per millisecond
Integration
Macro world
Nano world
Microsystems
Nano is an enabling technology; integration is the key!
Integrated Nanosystems
Data management
probing in space and time leads to a lot of data
x
y
z
t
(512x512) x 20 x 100 = 512 MB per cell
100 cells per condition
1 CD per cell
1 hard drive per experiment
SIMPLE ANALYSES
Where to meet nano …
security
– superior, lightweight materials
– advanced computing - unbreakable security, biometrics
– increased situational awareness - sensors
healthcare
– diagnostics – faster and cheaper, lab-on-chip, point-ofcare,
personal DNA mapping
– novel drugs – specific targeting, on-site local treatment
resources
– energy – effective utilization, alternatives – nanocatalysts
for solar generation of hydrogen from water
– water – purification and desalinization
Nano at internet
nano.gov … National Nanotechnology Initiative, USA
education.mrsec.wisc.edu/271.htm … Univ. Wisconsin video labs
nanobio.cz … our website with AFM resources (CZ)
nanocon.eu … international nano-conference in Brno
nanotechweb.org … IOP Institute of Physics
understandingnano.com …
imaginenano.info .. MathScience Innovation Center
nisenet.org … Nanoscal Informal Science Education
nano-ed.org … NSF-supported web community
trynano.org … IEEE
nanobio.cz … our website with AFM resources (CZ)
nanocon.eu … international nano-conference in Brno
nanometrologie.cz … Czech Metrological Institute (Gwydion)
Nano-books
D.S. Goodsell: Bionanotechnology Lessons from Nature, Wiley-Liss, 2004
E.S. Papazoglou, A. Parthasarathy: BioNanotechnology, Morgan & Claypool
Publishers, 2005
E. Gazit: Plenty of Room for Biology at the Bottom, Imperial College Press,
2007
to be continued …
Conclusions
modern science pushes the limits of
– creativity
– understanding
– technology
– information management
need
– ability to collect, store and retrieve large data sets
rapidly
– ability to analyze and reduce information to
comprehensible principles in real time
– ability to visualize and present conclusions and data
– ability to interpret in context of biological problem