Onofrio Annunziata, PhD , Texas Christian University
Diffusion of salt ions in water in the presence of proteins: Probing hydration of enzymes through the obstruction effect. Transport properties of saccharide-salt aqueous mixtures are important for basic research and applications in the biotechnological fields. We measured accurate NaCl main-term diffusion coefficients in aqueous lysozyme solutions at 25 ºC and pH 4.5 using the Gosting diffusiometer operated in its Rayleigh interferometric optical mode. The dependence of this diffusion coefficient on lysozyme concentration was examined using the obstruction-effect theory. Agreement between experimental results and theory is achieved if lysozyme proteins are treated as hydrated spheres with a hydration number of 240. Electrostatic interactions and common-ion effects due to lysozyme net charge at pH 4.5 do not contribute significantly to the behavior of the NaCl diffusion coefficient within our experimental range of salt concentrations (0.25-0.90 M). This work shows that the obstruction effect of proteins on small ions can be used to probe protein hydration, an important aspect of enzyme catalysis. This work has been submitted for publication to J. Chem. Eng. Data.
Polymeric aggregates with potential catalytic activity. We have previously shown that the morphology of protein aggregates is controlled by the presence/absence of phase transitions such as liquid-liquid phase separation (LLPS). Phase transitions are related to the nature and concentration of cosolvents. This concept was then extended to hydrophilic polymers such as polyvinyl alcohol (PVA). For PVA (100 kg/mol) in water in the presence of polyethylene glycol (8 kg/mol), LLPS was observed. However, in the presence of salts, PVA formed amorphous aggregates. Dynamic-light-scattering measurements revealed that salts lead to the formation of PVA mesoscopic aggregates with tunable size (from 20 nm to 500 nm) (September 2009-September 2010 period). Our objective is to use these PVA aggregates as scaffold for catalytically active molecules. We have conisfound that a few porphyrin molecules bind to PVA by performing spectrophotometric measurements and partitioning studies in the presence of PVA hydrogel. We believe that our investigation will lead to the preparation of catalytically active polymeric aggregates with properties similar to that of redox proteins such as cytochromes. In the September 2010-September 2011 period, we have finished our systematic characterization and one manuscript is currently in preparation.