Ligand-induced conformational changes in lactose repressor: a phosphorescence and ODMR study of single-tryptophan mutants.
Phosphorescence and optically detected magnetic resonance (ODMR) measurements are reported on four single-tryptophan mutants of lac repressor protein from Escherichia coli: H74W/Wless, W201Y, Y273W/Wless, and F293W/Wless, where Wless represents a protein background containing the double mutation W201Y/W220Y. The single-tryptophan residues are located in the protein core region, either in the monomer-monomer interface of the tetrameric protein or in the region of the inducer binding cleft. Inducer binding elicits large changes in the energy (0,0-band wavelength shifts) and zero-field splitting energies (ZFS) of the triplet states for each of the mutant proteins except W201Y which exhibits more modest effects. F293W/Wless exists in two distinguishable conformations, only one of which appears to be sensitive to the presence of inducer. These effects of inducer binding can be attributed to a conformational change that alters specific polar interactions that occur at each affected tryptophan site. Changes in the tryptophan triplet state indicator depend on the existence of specific polar interactions that are altered by local atomic relocations.
Ozarowski
A
A
Department of Chemistry, University of California, Davis 95616, USA.
Barry
J K
JK
Matthews
K S
KS
Maki
A H
AH
D019943:Amino Acid Substitution / Q000235:genetics; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics*; D004926:Escherichia coli / Q000737:chemistry / Q000235:genetics; D029968:Escherichia coli Proteins; D006639:Histidine / Q000235:genetics; D007544:Isopropyl Thiogalactoside / Q000737:chemistry; D056968:Lac Repressors; D007785:Lactose / Q000037:antagonists & inhibitors; D008024:Ligands; D008163:Luminescent Measurements; D046911:Macromolecular Substances; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D010649:Phenylalanine / Q000235:genetics; D011487:Protein Conformation; D012097:Repressor Proteins / Q000737:chemistry* / Q000235:genetics*; D014364:Tryptophan / Q000235:genetics*; D014443:Tyrosine / Q000235:genetics
D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D056968:Lac Repressors; C539064:LacI protein, E coli; D008024:Ligands; D046911:Macromolecular Substances; D012097:Repressor Proteins; D007544:Isopropyl Thiogalactoside; D014443:Tyrosine; D010649:Phenylalanine; D006639:Histidine; D014364:Tryptophan; D007785:Lactose