An essential intermediate in the folding of dihydrofolate reductase.
The folding of Escherichia coli dihydrofolate reductase was examined at pH 7.8 and 15 degrees C by using stopped-flow fluorescence and absorbance spectroscopies. The formation of a highly fluorescent intermediate occurs with relaxation times ranging between 142 and 343 msec, whereas stopped-flow absorbance spectroscopy using methotrexate binding assays shows a distinct lag phase during these time frames for the native state. The lag in absorbance kinetics and the lack of fast-track folding events indicate that the formation of this ensemble of intermediates is an obligatory step in the folding reaction.
Heidary
D K
DK
Department of Chemistry and Biochemistry, University of California, San Diego, La Jolla, CA 92093-0359, USA.
O'Neill
J C
JC
Roy
M
M
Jennings
P A
PA
D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology; D008727:Methotrexate / Q000378:metabolism; D008958:Models, Molecular; D011485:Protein Binding; D011487:Protein Conformation; D017510:Protein Folding; D013050:Spectrometry, Fluorescence; D013762:Tetrahydrofolate Dehydrogenase / Q000737:chemistry* / Q000378:metabolism
D001426:Bacterial Proteins; D013762:Tetrahydrofolate Dehydrogenase; D008727:Methotrexate