Biomedical Engineering Reference
In-Depth Information
both a long T (22 mer) and a long A (30 mer) tract. As the Figure 1.42 graph inset clearly
shows, by around a length of 4-8 bp the frequency/bp for A tracts from three different
eukaryotes has significantly increased relative to frequencies expected for random tract
occurrence in an organism comprised of random sequences of the same overall base com-
position. However, for
D. discoideum
sequences the relative frequency increase is remark-
able. Tracts of A above 40 bp length were observed, with two A tracts even greater than 50
bp, representing a frequency/bp that is 10
15
-fold above that expected to occur within ran-
dom sequence DNA of equivalent base composition (120,121). In this study, nearly as long
a tract distribution was observed within the genome of the (A
T)% base-rich organism
Plasmodium falciparum
(the malaria parasite). The
redundancy
built into the occurrence of
these homopolymer tracts has specific functions in the DNA of these organisms that is just
beginning to be elucidated. Some of the functions may result from the unusual structures
of these DNA tracts. Short A and T run homopolymer tracts (3-5 bp) are known to exhibit
helix axis bending of around 15-20° (122). In some biological situations, such as in the kin-
teoplast DNA of primitive eukaryotes, these tract bends are found in phase with the turn
of the helix at lengths of 10-11 bp, resulting in the large-scale macroscopic bending of long
stretches of DNA hundreds of base pairs in length (123). More commonly observed func-
tions for A and T homopolymer tracts at lengths of 10 bp or greater are the transcriptional
regulation of genes (124), where tract-specific recognition proteins bind the tracts and act
as promoters of specific gene transcription.
We have also observed that the longer A and T tracts (
8-10 bp) are not located ran-
domly in genomic DNA but are found on average to be spaced at preferred distances
apart. In the case of
D. discoideum
(125) and a few other eukaryotes (126), this spacing cor-
responds to the measured average spacing of the organism's nucleosomes, the repeating
nucleoprotein structural complex found in the chromosomes of all eukaryotic organisms.
This fact suggests that these homopolymer tracts may exist within specific cellular chro-
mosomal packaging environments in the cell as well as possess functions that are distinct
from the average DNA sequence found in every organism.
1.3.3
DNA Tertiary Structure Self-Assembly—Counterion Condensation Drives
Intramolecular DNA Collapse and Helps Determine Electrophoretic Mobilities
Self-assembly
is a property of DNA that is exhibited in a number of different ways.
Fundamentally, it underlies the double helical structure of DNA, via the molecular recog-
nition involved in complementary base pairing between single strands. This is taken
advantage of in countless ways in the molecular biology research laboratory and in the
biotechnology industry, through the use of DNA hybridization assays in various research
protocols. The DNA hybridization process has also been incorporated into the design of
biosensors. A common strategy is to hybridize a single-stranded sequence probe, often
immobilized onto a solid support, with an analyte single-stranded sequence from a sam-
ple where knowledge of the presence and sometimes concentration of the nucleic acid is
desired. This
self-assembly
hybridization strategy is
discriminating in its recognition
and sen-
sor function. The effect of increasing base mismatch level in imperfectly matched sequence
hybrids produces thermodynamically unstable hybrids that are not sensed. This feature is
typically taken advantage of in biosensor systems and provides them with a
discrimination
capability for detection of only the perfectly matched correct analyte sequence of interest.
As an example, biosensor systems incorporating a hybridization strategy can result in
detection of human pathogens possessing DNA sequences unique from our own.
Therefore, hybridization-based biosensors can be used to identify infections in patient
samples (127).
