An interface point‐mutation variant of triosephosphate isomerase is compactly folded and monomeric at low protein concentrations

FEBS Letters

Jaenicke, R.³  Wierenga, R.K.¹  Zeelen, J.Ph.¹  Borchert, T.V.¹  Callens, M.²  Minke, W.²  Schliebs, W.¹ 

[1] EMBL, Postfach 102209, D-69012 Heidelberg, Germany;Research Unit for Tropical Diseases, International Institute of Cellular and Molecular Pathology, Avenue Hippocrate 74, B1200, Brussels, Belgium;Institut für Biophysik und Physikalische Biochemie, Universität Regensburg, D-93040, Regensburg, Germany

关键词: Dimer;    Triosephosphate isomerase;    Interface;    Ultracentrifugation;    Monomer;    Point mutation;    Bis-Tris;    bis[2-hydroxyethyl]imino-tris[hydroxymethyl]-methane;    CD;    circular dichroism;    DHAP;    dihydroxyacetone phosphate;    DTT;    dithiothreitol;    EDTA;    ethylenediamine tetraacetic acid;    GAP;    d-glyceraldehyde-3-phosphate;    TGGE;    temperature gradient gel electrophoresis;    TIM;    triosephosphate isomerase (EC 5.3.1.1);    wtTIM;    wild-type trypanosomal triosephosphate isomerase;    2PG;    2-phosphoglycollate;   

DOI  :  10.1016/0014-5793(95)00586-X

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

Wild-type trypanosomal triosephosphate isomerase (wtTIM) is a very tight dimer. The interface residue His-47 of wtTIM has been mutated into an asparagine. Ultracentrifugation data show that this variant (H47N) only dimerises at protein concentrations above 3 mg/ml. H47N has been characterised at a protein concentration where it is predominantly a monomer. Circular dichroism measurements in the near-UV and far-UV show that this monomer is a compactly folded protein with secondary structure similar as in wtTIM. The thermal stability of the monomeric H47N is decreased compared to wtTIM: temperature gradient gel electrophoresis (TGGE) measurements give T _m-values of 41°C for wtTIM, whereas the T _m-value for the monomeric form of H47N is approximately 7°C lower.

【 授权许可】

Unknown