Differential dehydration effects on globular proteins and intrinsically disordered proteins during film formation

Abstract

Globular proteins composed of different secondary structures and fold types were examined by synchrotron radiation circular dichroism spectroscopy to determine the effects of dehydration on their secondary structures. They exhibited only minor changes upon removal of bulk water during film formation, contrary to previously reported studies of proteins dehydrated by lyophilization (where substantial loss of helical structure and gain in sheet structure was detected). This near lack of conformational change observed for globular proteins contrasts with intrinsically disordered proteins (IDPs) dried in the same manner: the IDPs, which have almost completely unordered structures in solution, exhibited increased amounts of regular (mostly helical) secondary structures when dehydrated, suggesting formation of new intra‐protein hydrogen bonds replacing solvent‐protein hydrogen bonds, in a process which may mimic interactions that occur when IDPs bind to partner molecules. This study has thus shown that the secondary structures of globular and intrinsically disordered proteins behave very differently upon dehydration, and that films are a potentially useful format for examining dehydrated soluble proteins and assessing IDPs structures.

The GRASP domain in Golgi Reassembly and Stacking Proteins: differences and similarities between lower and higher Eukaryotes

“…Another strategy to indirectly probe whether a protein presents an IDP-like behavior is by checking the effects of dehydration on its secondary structure, which can be monitored by CD or SRCD spectroscopies [50,51]. Unlike IDPs, globular, well-structured proteins exhibit only minor alterations by CD upon removal of bulk water during film formation [51].…”

CDtoolX, a downloadable software package for processing and analyses of circular dichroism spectroscopic data

“…Concentration errors can be resolved by scaling and conversion to DE. The HT signal can be used as a diagnostic tool for problems such as light scattering, bubble formation in temperature scans, and precise data cutoff points (wavelengths at which too little light reaching the instrument detector can be measured – corresponding to the lowest wavelength where the data is valid). Moreover, facile comparison between data collected on different instruments and conventional and SRCD beamlines can be made.…”

“…CDtool has been widely used as a processing and analysis tool in CD and SRCD studies on diverse samples, including proteins, nucleic acids and even chiral small molecules, as well as for examining samples in different physical forms such as monolayers, fibers, emulsions, and oriented samples . Some recent examples of its use in different applications include: protein folding/unfolding, thermal stability profiles, comparisons of wild type and mutant proteins, monitoring continuous flow dynamics measurements, identifying similarities in protein evolutionary relationships, comparisons of new spectra with downloaded Protein Circular Dichroism Data Bank (PCDDB) components, demonstrations of fidelity of folding of expressed proteins, instrumentation comparisons and calibrations, as well as “on‐the‐fly” displays, data processing and analyses at SRCD beamlines .…”

Structural and dynamic characterization of the Golgi Reassembly and Stacking Protein (GRASP) in solution

“…It has been shown that IDPs can go through a disorder-to-order transition after/before (induced fit/conformation selection model) interaction with a protein partner to strengthen the complex stability [204,205,206,207]. Moreover, it has been recently observed that both native globular proteins and IDPs have very different structural responses after total dehydration [208]. While in the former no significant structure rearrangement is observed, in the latter multiple disorder-to-order transitions are induced [208].…”

“…Moreover, it has been recently observed that both native globular proteins and IDPs have very different structural responses after total dehydration [208]. While in the former no significant structure rearrangement is observed, in the latter multiple disorder-to-order transitions are induced [208]. Therefore, we investigated whether protein dehydration could play a role in inducing a disorder-to-order transition in CnGRASP, a native molten globule-like protein that might behave as an intermediate between regular globular proteins and extended IDPs.…”