Denaturation and partial renaturation of a tightly tetramerized DsRed protein under mildly acidic conditions

FEBS Letters

Varfolomeyev, Sergey D.²  Verkhusha, Vladislav V.¹  Vrzheshch, Peter V.¹  Akovbian, Nina A.² 

[1] Center for Molecular Medicine, Moscow State University, Vorob'evy Gory, Moscow 119899, Russia;Department of Chemical Enzymology, Faculty of Chemistry, Moscow State University, Vorob'evy Gory, Moscow 119899, Russia

关键词: Red fluorescent protein;    pH dependence;    Denaturation;    Renaturation;    Tetramer;    Kinetics;    GFP;    green fluorescent protein;    CD;    circular dichroism;   

DOI  :  10.1016/S0014-5793(00)02344-9

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

The red fluorescent protein, DsRed, recently cloned from coral Discosoma sp. has one of the longest fluorescence waves and one of the most complex absorbance spectra among the family of fluorescent proteins. In this work we found that with time DsRed fluorescence decreases under mildly acidic conditions (pH 4.0–4.8) in a pH-dependent manner, and this fluorescence inactivation could be partially recovered by subsequent re-alkalization. The DsRed absorbance and circular dichroism spectra under these conditions revealed that the fluorescence changes were caused by denaturation followed by partial renaturation of the protein. Further, analytical ultracentrifugation determined that native DsRed formed a tight tetramer under various native conditions. Quantitative analysis of the data showed that several distinct states of protein exist during the fluorescence inactivation and recovery, and the inactivation of fluorescence can be caused by protonation of a single ionogenic group in each monomer of DsRed tetramer.

【 授权许可】

Unknown