|
Scientists simulate DNA interacting with an engineered protein. The system may slow DNA strands travelling through pores enough to read a patient’s individual genome. Credit: of Aleksei Aksimentiev
|
The Human Genome Project paved the way for
genomics, the study of an organism's genome. Personalized genomics can
establish the relationship between DNA sequence variations among individuals
and their health conditions and responses to drugs and treatments. To make
genome sequencing a routine procedure, however, the time must be reduced to
less than a day and the cost to less than $1,000—a feat not possible with current
knowledge and technologies.
In 2008, a research team led by Aleksei
Aksimentiev, assistant professor in the physics department at the Univ. of Illinois-Urbana-Champaign, began a
project to create machines for personal genome sequencing that will be more
accessible to hospitals. Using Oak Ridge National Laboratory's Jaguar, one of
the world's fastest supercomputers, Aksimentiev and his team is developing a
nanopore approach, which promises a drastic reduction in time and costs for DNA
sequencing. Their research reveals the shape of DNA moving through a single
nanopore—a protein pore a billionth of a meter wide that traverses a membrane.
As the DNA passes through the pore, the sequence of nucleotides is read by a
detector.
"The main obstacle of sequencing using
the older generations of biological and synthetic nanopores was the inability
to identify the DNA sequence to single-nucleotide resolution," Aksimentiev
said. "The nucleotides passed too quickly through the nanopore for scientists
to sequence the DNA."
Aksimentiev's group uses the nanopore MspA,
an engineered protein. Its sequence must be altered to bind more strongly to
the moving DNA strand. MspA is an ideal platform for sequencing DNA because
scientists can now measure dams in the pore, which could slow DNA's journey
through the protein. Altering the MspA protein to optimize dams is both
time-consuming and costly in a laboratory but simple on a computer. For
instance, to alter the protein in any way, scientists must determine whether
the particular mutation they introduce is stable and if the idea is reasonable.
Therefore, the scientists first simulate MspA to decide on a mutation to induce
and to test high-risk ideas before implementing them in an experiment.
The research team uses the code NAMD, which
calculates minimum energy states of atoms in a large biomolecular system and is
an indicator of what shapes the molecules would be most comfortable assuming.
The team first builds a model of the MspA protein submerged in a lipid bilayer
and electrolyte solution. A DNA strand of a desired nucleotide sequence is then
threaded through the MspA nanopore. Next the scientists simulate the effect of
an electric field driving ions and DNA through the MspA nanopore. The
simulation employs molecular dynamics, or calculations of the motion of each
atom in a molecular system following the physical laws of nature, to mimic the
experimental system. The simulations' results can be directly compared to ones
from experiments because both approaches measure the ionic current, according
to Aksimentiev. By knowing the positions of each DNA atom and ion, scientists
gain an advantage—they can optimize nanopore sequencing using a rational design
to produce a pore that hugs to the DNA more tightly, slowing the molecule's
journey through the pore to a speed allowing single-nucleotide resolution.
The sequencing work is funded by the
National Human Genome Research Institute of the National Institutes of Health.
The project's method development is funded in part by the National Science
Foundation. Collaborators with the project include two experimental groups: one
led by Jens Gundlach at the Univ. of Washington-Seattle and the other by Michael
Niederweis at the Univ. of Alabama-Birmingham.
The research received 10 million processor
hours on Jaguar through the Innovative and Novel Computational Impact on Theory
and Experiment, or INCITE, program, which awards considerable allocations on
some of the world's most powerful supercomputers to projects addressing grand
challenges in science and engineering. With the INCITE allocation, the scientists
were able to reproduce the dams in the MspA nanopore for the type of DNA
nucleotides confined to it, slowing down the sequence movement through the
nanopore.
"We have carried out a pilot study on
several variants of the MspA nanopore and observed considerable reduction of
the DNA strand speed," Aksimentiev said. "These very preliminary
results suggest that achieving a 100-fold reduction of DNA velocity, which
should be sufficient to read out the DNA sequence with single-nucleotide resolution,
is within reach. Future studies will be directed toward this goal."
The team hopes to achieve this
project's objective by 2013 and plans to pursue a number of exciting spin-off
projects, Aksimentiev said. The ability to make genome sequencing affordable
will enable such programs as the Cancer Genome Project, which characterizes DNA
mutations in cancer cells in various tissues throughout all stages of cancer
development.
SOURCE
