The small angle X-ray scattering (SAXS) technique allows for the characterization of the size and shape of electron density inhomogeneities in matter from the nanometer to the micrometer scale. The experimental setup for these investigations is usually flexible enough to be easily applied to many samples in which the effect of changes in the chemical composition or preparation protocols can be probed, and the response of the system to stimuli can also be studied. The investigation of structure by means of SAXS under varying conditions can help elucidating the underlying mechanism of complex processes and linking structure to function.
The aim of the proposal is to provide machine time of the SAXSLab Sapienza for relatively time-consuming measurements of protein, peptide and self-assembled nanocarriers in solution under varying conditions, in order to answer questions posed by investigations in research topics related to protein functionality and the design of self-assembled structures:
a) Hydration associated with drug release in albumin-based non-covalent drug delivery systems.
b) Molecular aspects of protein oxidation and recovery.
c) Characterization of the structures formed by the self-assembly of peptide and peptide-amphiphiles and their inter-particle interactions.
d) Evaluation of the nanostructures of drug-loaded formulations based on amphiphilic molecules and copolymers.
We expect that experiments at SAXSLab Sapienza can provide advancements in research topics related to protein functionality and the design of self-assembled structures.
a) Hydration associated with drug release in albumin-based non-covalent drug delivery systems.
SAXS experiments at SAXSLab Sapienza will be useful to optimize the experimental conditions for the neutron beamtime session and to provide complementary information. Indeed, comparison of the scattered intensity and size determined from SAXS and SANS with different scattering contrast are fundamental for studying protein hydration [28]. At SAXSLab Sapienza it will be possible to study both low and high concentration samples at small and wide scattering angles, to monitor both the overall size (which include the hydration layer) and inter-particle interactions (at small angles) and changes in the wide angle scattering which also contains sub-nanometer scale information on solvation layers. Studies on albumin-drug complexes are very numerous in the literature, but insights from scattering techniques, and in particular studies addressing the coupling between water preferential interactions and ligand binding and release have a notable novelty content.
b) Molecular aspects of protein oxidation and recovery
Understanding the molecular basis of the protein resistance to chemical damage, or, on the other hand, of the processes leading to the formation of insoluble precipitates as a consequence of these oxidative modifications, can be relevant for understanding physiological phenomena taking place in conditions of chronical inflammation and oxidative stress. In addition, this knowledge can be also useful to prevent inactivation of useful enzymes or protein carriers in applications.
Studies on protein oxidation often neglect the specific effect of every step of the analysis protocols on the original situation that had to be characterized, and this is particularly problematic in the case of relatively unstable oxidant-induced modifications. We would like to demonstrate also with the use of SAXS measurements that the addition of reductants to "scavenge excess unreacted oxidant" in oxidation protocols for study purposes can determine a recovery from the protein modifications being studied.
c) Characterization of the structures formed by the self-assembly of peptide and peptide-amphiphiles and their inter-particle interactions.
The nanostructures formed by peptide and peptide-based amphiphiles have been intensively characterized in the last years. However systematic studies as a function of the concentration and solvent properties are rarely engaged and we forecast these can help elucidate still unclear phenomena in the inter-aggregates interactions which are of interest for both fundamental understanding of interaction potentials of complex colloids and applications due their impact on rheological and stability properties.
In the field of peptide-based amphiphiles, the use of building blocks with D- instead of L-aminoacids
d) Evaluation of the nanostructures of drug-loaded formulations based on amphiphilic molecules and copolymers.
Characterization with the SAXS technique can increase the reliability of the systems proposed for potential applications in appropriate solution conditions and consequently the impact in the scientific community.