3 research outputs found
Determination of Peptide<b>–</b>Surface Adsorption Free Energy for Material Surfaces Not Conducive to SPR or QCM using AFM
The interactions between peptides and proteins with material
surfaces
are of primary importance in many areas of biotechnology. While surface
plasmon resonance spectroscopy (SPR) and quartz crystal microbalance
(QCM) methods have proven to be very useful in measuring fundamental
properties characterizing adsorption behavior, such as the free energy
of adsorption for peptide–surface interactions, these methods
are largely restricted to use for materials that can readily form
nanoscale-thick films over the respective sensor surfaces. Many materials
including most polymers, ceramics, and inorganic glasses, however,
are not readily suitable for use with SPR or QCM methods. To overcome
these limitations, we recently showed that desorption forces (<i>F</i><sub>des</sub>) obtained using a standardized AFM method
linearly correlate to standard-state adsorption free energy values
(Δ<i>G°</i><sup><i></i></sup><sub>ads</sub>) measured from SPR in phosphate buffered saline (PBS: phosphate
buffered 140 mM NaCl, pH 7.4). This approach thus provides a means
to determine Δ<i>G°</i><sup><i></i></sup><sub>ads</sub> for peptide adsorption using AFM that can be
applied to any flat material surface. In this present study, we investigated
the <i>F</i><sub>des</sub>–Δ<i>G°</i><sup><i></i></sup><sub>ads</sub> correlation between AFM
and SPR data in PBS for a much broader range of systems including
eight different types of peptides on a set of eight different alkanethiol
self-assembled monolayer (SAM) surfaces. The resulting correlation
was then used to estimate Δ<i>G°</i><sup><i></i></sup><sub>ads</sub> from <i>F</i><sub>des</sub> determined by AFM for selected bulk polymer and glass/ceramic materials
such as polyÂ(methyl methacrylate) (PMMA), high-density polyethylene
(HDPE), fused silica glass, and a quartz (100) surface. The results
of these studies support our previous findings regarding the strong
correlation between <i>F</i><sub>des</sub> measured by AFM
and Δ<i>G°</i><sup><i></i></sup><sub>ads</sub> determined by SPR, and provides a means to estimate Δ<i>G°</i><sup><i></i></sup><sub>ads</sub> for peptide
adsorption on macroscopically thick samples of materials that are
not conducive for use with SPR or QCM
Structure of Hydrated Poly(d,l-lactic acid) Studied with X-ray Diffraction and Molecular Simulation Methods
The effect of hydration on the molecular structure of
amorphous
polyÂ(d,l-lactic acid) (PDLLA) with 50:50 L-to-D
ratio has been studied by combining experiments with molecular simulations.
X-ray diffraction measurements revealed significant changes upon hydration
in the structure functions of the copolymer. Large changes in the
structure functions at ∼10 days of incubation coincided with
the large increase in the water uptake from ∼1 to ∼40%
and the formation of voids in the film. Computer modeling based on
the recently developed TIGER2/TIGER3 mixed sampling scheme was used
to interpret these changes by efficiently equilibrating both dry and
hydrated models of PDLLA. Realistic models of bulk amorphous PDLLA
structure were generated as demonstrated by close agreement between
the calculated and the experimental structure functions. These molecular
simulations were used to identify the interactions between water and
the polymer at the atomic level including the change of positional
order between atoms in the polymer due to hydration. Changes in the
partial O–O structure functions, about 95% of which were due
to water–polymer interactions, were apparent in the radial
distribution functions. These changes, and somewhat smaller changes
in the C–C and C–O partial structure functions, clearly
demonstrated the ability of the model to capture the hydrogen-bonding
interactions between water and the polymer, with the probability of
water forming hydrogen bonds with the carbonyl oxygen of the ester
group being about 4 times higher than with its ether oxygen
Adsorption-Induced Changes in Ribonuclease A Structure and Enzymatic Activity on Solid Surfaces
Ribonuclease
A (RNase A) is a small globular enzyme that lyses
RNA. The remarkable solution stability of its structure and enzymatic
activity has led to its investigation to develop a new class of drugs
for cancer chemotherapeutics. However, the successful clinical application
of RNase A has been reported to be limited by insufficient stability
and loss of enzymatic activity when it was coupled with a biomaterial
carrier for drug delivery. The objective of this study was to characterize
the structural stability and enzymatic activity of RNase A when it
was adsorbed on different surface chemistries (represented by fused
silica glass, high-density polyethylene, and polyÂ(methyl-methacrylate)).
Changes in protein structure were measured by circular dichroism,
amino acid labeling with mass spectrometry, and in vitro assays of
its enzymatic activity. Our results indicated that the process of
adsorption caused RNase A to undergo a substantial degree of unfolding
with significant differences in its adsorbed structure on each material
surface. Adsorption caused RNase A to lose about 60% of its native-state
enzymatic activity independent of the material on which it was adsorbed.
These results indicate that the native-state structure of RNase A
is greatly altered when it is adsorbed on a wide range of surface
chemistries, especially at the catalytic site. Therefore, drug delivery
systems must focus on retaining the native structure of RNase A in
order to maintain a high level of enzymatic activity for applications
such as antitumor chemotherapy