Effects of macromolecular crowding on the structural stability of human α-lactalbumin.
The folding of protein, an important process for protein to fulfill normal functions, takes place in crowded physiological environments. α-Lactalbumin, as a model system for protein-folding studies, has been used extensively because it can form stable molten globule states under a range of conditions. Here we report that the crowding agents Ficoll 70, dextran 70, and polyethylene glycol (PEG) 2000 have different effects on the structural stability of human α-lactalbumin (HLA) represented by the transition to a molten globule state: dextran 70 dramatically enhances the thermal stability of Ca(2+)-depleted HLA (apo-HLA) and Ficoll 70 enhances the thermal stability of apo-HLA to some extent, while PEG 2000 significantly decreases the thermal stability of apo-HLA. Ficoll 70 and dextran 70 have no obvious effects on trypsin degradation of apo-HLA but PEG 2000 accelerates apo-HLA degradation by trypsin and destabilizes the native conformation of apo-HLA. Furthermore, no interaction is observed between apo-HLA and Ficoll 70 or dextran 70, but a weak, non-specific interaction between the apo form of the protein and PEG 2000 is detected, and such a weak, non-specific interaction could overcome the excluded-volume effect of PEG 2000. Our data are consistent with the results of protein stability studies in cells and suggest that stabilizing excluded-volume effects of crowding agents can be ameliorated by non-specific interactions between proteins and crowders.
State Key Laboratory of Virology, College of Life Sciences,  Wuhan University, Wuhan 430072,  China.
De-Lin
DL
Zhang
Ling-Jia
LJ
Wu
Jie
J
Chen
Yi
Y
Liang
D002118:Calcium / Q000737:chemistry; D002151:Calorimetry / Q000379:methods; D002942:Circular Dichroism; D006801:Humans; D007768:Lactalbumin / Q000737:chemistry*; D046911:Macromolecular Substances; D008968:Molecular Conformation; D011092:Polyethylene Glycols / Q000737:chemistry; D011487:Protein Conformation; D011489:Protein Denaturation; D017510:Protein Folding; D011506:Proteins / Q000737:chemistry; D013050:Spectrometry, Fluorescence / Q000379:methods; D013816:Thermodynamics; D014357:Trypsin / Q000737:chemistry
D046911:Macromolecular Substances; D011506:Proteins; D011092:Polyethylene Glycols; D007768:Lactalbumin; D014357:Trypsin; D002118:Calcium