Authors:
SP Williams, AM Fulton, KM Brindle
Journal name: 
Biochemistry
Citation info: 
32(18):4895-4902
Abstract: 
Yeast phosphoglycerate kinase was selectively fluorine-labeled in vivo by inducing enzyme synthesis in stationary phase cells in the presence of 5-fluorotryptophan. Inducible expression was obtained using a galactose-inducible expression vector containing the yeast phosphoglycerate kinase coding sequence. 19F NMR measurements on intact cells showed two resolved resonances, from the two tryptophan residues in the protein, which underwent reversible changes in chemical shift under different metabolic conditions. Measurements in vitro showed that the difference in the chemical shifts of these two resonances was dependent on the adenine nucleotide concentration, in particular the MgADP concentration. A comparison of the spectra obtained in vitro with those obtained from the intact cell indicated that in glucose-fed cells the cytosolic free MgADP concentration was less than 50 microM, which is significantly lower than the concentrations measured in whole-cell extracts.
DOI: 
http://doi.org/10.1021/bi00069a026
Research group: 
Brindle Group
E-pub date: 
30 Apr 1993
Users with this publication listed: 
Kevin Brindle