title: Peculiarities of the Super-Folder GFP Folding in a Crowded Milieu.
abstract: The natural cellular milieu is crowded by large quantities of various biological macromolecules. This complex environment is characterized by a limited amount of unoccupied space, limited amounts of free water, and changed solvent properties. Obviously, such a tightly packed cellular environment is poorly mimicked by traditional physiological conditions, where low concentrations of a protein of interest are analyzed in slightly salted aqueous solutions. An alternative is given by the use of a model crowded milieu, where a protein of interest is immersed in a solution containing high concentrations of various polymers that serve as model crowding agents. An expected outcome of the presence of such macromolecular crowding agents is their ability to increase conformational stability of a globular protein due to the excluded volume effects. In line with this hypothesis, the behavior of a query protein should be affected by the hydrodynamic size and concentration of an inert crowder (i.e., an agent that does not interact with the protein), whereas the chemical nature of a macromolecular crowder should not play a role in its ability to modulate conformational properties. In this study, the effects of different crowding agents (polyethylene glycols (PEGs) of various molecular masses (PEG-600, PEG-8000, and PEG-12000), Dextran-70, and Ficoll-70) on the spectral properties and unfolding-refolding processes of the super-folder green fluorescent protein (sfGFP) were investigated. sfGFP is differently affected by different crowders, suggesting that, in addition to the expected excluded volume effects, there are some changes in the solvent properties.
authors: Laboratory of Structural Dynamics, Stability and Folding of Proteins, Institute of Cytology, Russian Academy of Sciences, 4 Tikhoretsky Ave., St. Petersburg 194064, Russia. lvs@incras.ru.
authors: Olesya V
authors: OV
authors: Stepanenko
authors: Laboratory of Structural Dynamics, Stability and Folding of Proteins, Institute of Cytology, Russian Academy of Sciences, 4 Tikhoretsky Ave., St. Petersburg 194064, Russia. sov@incras.ru.
authors: Olga V
authors: OV
authors: Stepanenko
authors: Laboratory of Structural Dynamics, Stability and Folding of Proteins, Institute of Cytology, Russian Academy of Sciences, 4 Tikhoretsky Ave., St. Petersburg 194064, Russia. imk@incras.ru.
authors: Irina M
authors: IM
authors: Kuznetsova
authors: Laboratory of Structural Dynamics, Stability and Folding of Proteins, Institute of Cytology, Russian Academy of Sciences, 4 Tikhoretsky Ave., St. Petersburg 194064, Russia. vuversky@health.usf.edu.|Department of Molecular Medicine and USF Health Byrd Alzheimer's Research Institute, Morsani College of Medicine, University of South Florida, 12901 Bruce B. Downs Blvd. MDC07, Tampa, FL 33612, USA. vuversky@health.usf.edu.
authors: Vladimir N
authors: VN
authors: Uversky
authors: Laboratory of Structural Dynamics, Stability and Folding of Proteins, Institute of Cytology, Russian Academy of Sciences, 4 Tikhoretsky Ave., St. Petersburg 194064, Russia. kkt@incras.ru.|Institute of Physics, Nanotechnology and Telecommunications, Peter the Great St. Petersburg State Polytechnic University, 29 Polytechnicheskaya st., St. Petersburg 195251, Russia. kkt@incras.ru.
authors: Konstantin K
authors: KK
authors: Turoverov
mesh_terms: D060833:Cellular Microenvironment / Q000235:genetics*; D049452:Green Fluorescent Proteins / Q000737:chemistry* / Q000235:genetics; D008970:Molecular Weight; D011092:Polyethylene Glycols / Q000737:chemistry; D011108:Polymers / Q000737:chemistry; D011487:Protein Conformation; D017510:Protein Folding; D012996:Solutions / Q000737:chemistry; D012997:Solvents / Q000737:chemistry; D014867:Water / Q000737:chemistry
chemical_list: D011108:Polymers; D012996:Solutions; D012997:Solvents; D014867:Water; D049452:Green Fluorescent Proteins; D011092:Polyethylene Glycols
keywords: conformational stability; crowded milieu; excluded volume effect; protein folding; solvent properties; super-folder GFP