Denaturant-induced expansion and compaction of a multi-domain protein: IgG.
It is generally believed that unfolded or denatured proteins show random-coil statistics and hence their radius of gyration simply scales with solvent quality (or concentration of denaturant). Indeed, nearly all proteins studied thus far have been shown to undergo a gradual and continuous expansion with increasing concentration of denaturant. Here, we use fluorescence correlation spectroscopy (FCS) to show that while protein A, a multi-domain and predominantly helical protein, expands gradually and continuously with increasing concentration of guanidine hydrochloride (GdnHCl), the F(ab')2 fragment of goat anti-rabbit antibody IgG, a multi-subunit all beta-sheet protein does not show such continuous expansion behavior. Instead, it first expands and then contracts with increasing concentration of GdnHCl. Even more striking is the fact that the hydrodynamic radius of the most expanded F(ab')2 ensemble, observed at 3-4 M GdnHCl, is approximately 3.6 times that of the native protein. Further FCS measurements involving urea and NaCl show that the unusually expanded F(ab')2 conformations might be due to electrostatic repulsions. Taken together, these results suggest that specific interactions need to be considered while assessing the conformational and statistical properties of unfolded proteins, particularly under conditions of low solvent quality.
Guo
Lin
L
Department of Chemistry, School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA.
Chowdhury
Pramit
P
Glasscock
Julie M
JM
Gai
Feng
F
D000818:Animals; D004058:Diffusion / Q000187:drug effects; D006041:Goats; D019791:Guanidine / Q000494:pharmacology*; D007140:Immunoglobulin Fab Fragments / Q000737:chemistry; D007074:Immunoglobulin G / Q000737:chemistry* / Q000378:metabolism*; D011489:Protein Denaturation / Q000187:drug effects; D017510:Protein Folding / Q000187:drug effects*; D017434:Protein Structure, Tertiary; D013997:Time Factors
D007140:Immunoglobulin Fab Fragments; D007074:Immunoglobulin G; D019791:Guanidine