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Biomolecular systems (523-531)
COLL
523: First experiments on electro-optic properties
of biopolymers
Henri C. Benoit, Institut Charles Sadron, 6 rue Boussingault, 67083
Strasbourg Cedex, France, benoit@cerbere.u-strasbg.fr
Abstract
A historical account is given of the first electrical birefringence (EB)
studies of biopolymers 50 years ago. My thesis project on EB of classical
polymer solutions resulted in practically the same effects for polymers
and monomeric units, and very high voltage was required. This led me to
look at rigid molecules like DNA and tobacco mosaic virus (TMV). Due to
the high conductivity of the solutions, I decided to use pulses. The
observed optical signal was asymmetric and deformed by the non-negligible
value of the rotary diffusion constant D. Hence, I developed the
theory of this effect. It shows that, regardless of the mechanism of
orientation, the decay of the signal is proportional to exp(-6Dt).
Very high values of D were found for both TMV (as expected) and
DNA. For the latter, the result was difficult to explain as the double
helix structure was not yet known at that time. Full
paper
COLL
524: Reversing-pulse electric birefringence of
multicomponent systems: The case of negative and positive mixed signals of
DNA in ionic solutions
Kiwamu Yamaoka, Faculty of Science, Hiroshima University, 1-3-1
Kagamiyama, (home: 5-chome, 2-2-805, Takaya Takamigaoka, Higashi-Hioshima
739-2115), Higashi-Hiroshima 739-8526, Japan, Fax: 0824-34-2920, yamaoka@hiroshima-u.ac.jp
Abstract
This presentation contains two parts on reversing-pulse electric
birefringence (RPEB) signal patterns. The first is the theoretical
calculation for two rod-like models coexisting in solution. RPEB curves
were calculated with a theory for ion-fluctuation dipole moment along
longitudinal direction with various electric and optical parameters
assigned to the models. Often a net decay curve appears faster than that
of either component, crossing the baseline only if a positive and a
negative EB component are present. The second part is the analysis of
experimental signals of sonicated and column-fractionated three calf
thymus DNA samples, measured at 7 °C in Na+ or Mg2+
solutions. While MgDNA samples mostly show normal RPEB signals, NaDNA
samples exhibit complex patterns, mixed with negative- and positive-going
profiles. By analyzing abnormal signals with calculations, a dip in the
reverse process with a larger relaxation time is confirmed for the
positive-going signal.
COLL
525: DNA oligomers studied by the optical Kerr
effect
Sergio R. Aragon, and Martin Perez, Department of Chemistry &
Biochemistry, San Francisco State University, 1600 Holloway Avenue, San
Francisco, CA 94132, aragons@sfsu.edu
Abstract
The optical Kerr effect shows promise as a technique for the analysis of
nucleic acid structure despite the earlier negative result of Norden and
collaborators (J. Phys. Chem. 91, 1957-60, 1987). A high power Nd:YAG (l=532nm)
pulsed laser was used to induce readily measurable transient birefringence
in an aqueous solution of short DNA oligomers. The birefringence was
probed with an Argon ion laser (l=488nm),
utilizing suppression of the solvent birefringence. In our experiment it
is possible to measure the rotational relaxation rates as well as the
birefringence of nucleic acids. The relaxation rates for several short
oligomers were measured and compared well to those from standard
hydrodynamics calculations, and agreed with published dynamic light
scattering measurements. With appropriate physical model for the
relaxation processes and the polarizability anisotropy of the molecule, it
may be possible to quantitatively determine the tertiary structure of a
nucleic acid.
COLL
526: Using transient electric birefringence to
measure the flexibility and static bending of DNA restriction fragments
Nancy C. Stellwagen, Yongjun Lu, and Brock Weers, Department of
Biochemistry, University of Iowa, Iowa City, IA 52242, Fax: 319-335-9570,
nancy-stellwagen@uiowa.edu
Abstract
Transient electric birefringence (TEB) was used to measure the persistence
length of fifteen DNA restriction fragments ranging from 79-789 bp in
length, in three low ionic strength buffers and at temperatures ranging
from 4º - 43º C. The restriction fragments are not curved or bent. The
persistence length and hydrodynamic radius were determined from a global
least squares fit of the terminal relaxation times to various hydrodynamic
equations. TEB was also used to determine the bend angle at the M13 origin
of replication by comparing the terminal relaxation times of bent and
unbent DNA fragments of the same size. Comparison of the terminal
relaxation times of four overlapping fragments of different sizes with
theoretical curves calculated for once-broken rods or wormlike coils
allowed the position of the bend to be located within ± 2 bp and the
magnitude of the bend angle to be determined without prior assumptions.
COLL
527: Significant differences are observed between
gel mobility anomalies and TEB relaxation times for curved DNA molecules
Yongjun Lu, Brock Weers, and Nancy C. Stellwagen, Department of
Biochemistry, University of Iowa, Newtown Street, Iowa City, IA 52242,
Fax: 319-335-9570, yongjun-lu@uiowa.edu
Abstract
Bent DNA fragments migrate anomalously slowly in polyacrylamide gels
because of additional friction with the gel matrix. DNA bending also
results in anomalously short transient electric birefringence (TEB)
rotational relaxation times (t) in
solution, due to the decreased length of the bent fragment. An apparent
bend of 45 ± 3° in simian virus 40 was found at ~ 1922-bp, flanked by A6-
and T7-tracts on either side. To check the sequence dependence
of the observed bend angles, fragments of ~ 220 bp were constructed by
inserting different linkers between the A6- and T7-tracts
in the target sequence in SV40. The insertion of a 25-26 bp spacer or two
14 bp spacers made the newly constructed fragment perform like an unbent
one (TEB-t measurements) while the
gel mobility anomalies continued to be observed. The results were analyzed
based on DNA anisotropic flexibility and helical phasing of the flanking
A- and T-tracts.
COLL
528: Electric dipole moments of bacteria:
Electro-optic measurements of the permanent and induced dipole moments of Escherichia
coli and Pseudomonas aeruginosa 11B
Stoyl P. Stoylov, Institute of Physical Chemistry, Bulgarian
Academy of Sciences, 1113 Sofia, Bulgaria
Abstract
The dipole moments of the bacteria were measured by electric light
scattering (ELS) (Stoylov et al. in Sofia and Jennings and Morris in
Uxbridge) and dichroism in rotating electric field (DREF). Some essential
contradictions appeared between the values and direction of the permanent
dipole moments, and also for the value of the induced dipole moment. It
seems that the communicated values for the electric polarizability of
bacteria are rather low compared to that generally found for
nanoparticles, if one expects that its strong (quadratic or cubic)
dependence on the size holds. Strong effects of surfactant, alcohol and
ether on the electric dipole moments of bacteria have been found. It seems
that a correlation exists between the bacteria's electric polarizability
and their viability. The possible connection between bacteria's dipole
moments and bacterial adhesion as well as other bacterial properties will
be discussed.
COLL
529: Electrokinetic mobility of short DNA
oligomers
Nancy C. Stellwagen, Department of Biochemistry, University of
Iowa, Iowa City, IA 52242, Fax: 319-335-9570, nancy-stellwagen@uiowa.edu,
and Alexandar M. Zhivkov, Institute of Physical Chemistry, Bulgarian
Academy of Sciences
Abstract
The electrophoretic mobility of single-stranded DNA oligomers containing
1-20 nucleotides have been measured by capillary electrophoresis in Tris-acetate
and isoelectric histidine buffers. The free solution mobility increases
with increasing molecular size until reaching a plateau and leveling off
for oligomers containing 10 or more nucleotides. This behavior is
unexpected because counterion condensation should be less complete near
the ends of small oligomers, according to Manning's theory of strong
polyelectrolytes. As a result, the electrophoretic mobility should
decrease with increasing DNA molecular weight, if the ratio of effective
electric charge to friction coefficient is constant. The observed
discrepancy can be explained by the higher hydration of shorter oligomers
caused by their larger effective electric charge. DNA-histidine complexes
will also be discussed. This work is supported by NATO Collaborative
Linkage Grant LST.CLG.978851.
COLL
530: Electro-optical study of plastocyanin
adsorbed on b-ferric
hydroxide particles
Nikolai Tuparev1, Ivana B. Petkanchin2,
Antony Donchev1, Mitko Dimitrov1, and Stefka G.
Taneva1. (1) Institute of Biophysics, Bulgarian Academy of
Sciences, Acad. G. Bontchev str., bl. 21, Sofia 1113, Bulgaria, niki@obzor.bio21.bas.bg,
(2) Institute of Physical Chemistry, Bulgarian Academy of Sciences
Abstract
The blue copper-containing protein plastocyanin (PC) acts as an electron
carrier in the light reaction of photosynthesis. It is believed that its
electrical state would play an important role in the photosynthetic
processes. The adsorption of two types of plastocyanin, PCa and
PCb, in oxidized form, on b-ferric
hydroxide particles is studied electro-optically. PCb has a
larger net negative charge than PCa. Electrostatic interactions
lead to the adsorption of PC molecules on positively charged oxide
particles. A decrease in the electric polarizability is found upon
adsorption of plastocyanin on the hydroxide particles when the pH is
varied from 4 to 10, with the effect being more pronounced for PCb.
The results demonstrate that electro-optics can be applied to study the
electrical state of small protein molecules by adsorbing them on model
anisometric particles.
COLL
531: Using capillary electrophoresis to measure
the electrostatic shielding of DNA in various buffer/salt solutions
Nancy C. Stellwagen, and Earle Stellwagen, Department of
Biochemistry, University of Iowa, Iowa City, IA 52242, Fax: 319-335-9570,
nancy-stellwagen@uiowa.edu
Abstract
Free solution mobility of 20-bp double-stranded DNA oligomer was measured
in diethylmalonate (DM) and Tris-acetate buffers, with and without added
NaCl or TrisCl. The mobility decreases linearly with the logarithm of
ionic strength when increasing the buffer concentration, and also when
NaCl is added to NaDM buffer or TrisCl is added to Tris DM buffer.
Non-linear effects are observed when the counterion in the added salt
differs from the counterion in the buffer. The dependence of the mobility
on ionic strength cannot be predicted using the Henry, Debye-Huckel or
Pitts equations for electrophoresis. However, the mobilities observed in
all buffer and buffer/salt solutions can be predicted within ~ 20 % from
the Manning equation for electrophoresis, using no adjustable parameters.
The results suggest that the electrostatic shielding of DNA is determined
not only by the relative concentrations of the various ions in the
solution, but also by their equivalent conductivities.
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