Small-angle X-ray scattering study of the kinetics of light-dark transition in a LOV protein

• Published in: PloS one Vol. 13; no. 7; p. e0200746
• Main Authors: Röllen, Katrin, Granzin, Joachim, Batra-Safferling, Renu, Stadler, Andreas Maximilian
• Format: Journal Article
• Language: English
• Published: United States Public Library of Science 16.07.2018; Public Library of Science (PLoS)
• Subjects: Analysis; Archaea; Bacterial Proteins - metabolism; Biochemistry; Biology and Life Sciences; Chromophores; Crystallography; Crystallography, X-Ray; Flavin; Flavin Mononucleotide - chemistry; Flavoproteins; Flavoproteins - metabolism; Fungi; Kinetics; Light; LOV protein; NMR; Nuclear magnetic resonance; Oxygen; Oxygen - metabolism; Photoreceptors; Photoreceptors, Microbial - metabolism; Physical Sciences; Protein Domains; Protein Structure, Secondary; Proteins; Pseudomonas putida; Pseudomonas putida - metabolism; Recovery; Recovery time; Relaxation time; Research and Analysis Methods; Scattering, Small Angle; Signal transduction; Small angle X ray scattering; Social Sciences; Spectrophotometry, Ultraviolet; Spectroscopy; Spectrum analysis; Structural dynamics; Time measurement; Ultraviolet-visible spectroscopy; X ray scattering; X-Ray Diffraction; X-ray spectroscopy; Germany
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0200746

Light, oxygen, voltage (LOV) photoreceptors consist of conserved photo-responsive domains in bacteria, archaea, plants and fungi, and detect blue-light via a flavin cofactor. We investigated the blue-light induced conformational transition of the dimeric photoreceptor PpSB1-LOV-R66I from Pseudomonas putida in solution by using small-angle X-ray scattering (SAXS). SAXS experiments of the fully populated light- and dark-states under steady-state conditions revealed significant structural differences between the two states that are in agreement with the known structures determined by crystallography. We followed the transition from the light- to the dark-state by using SAXS measurements in real-time. A two-state model based on the light- and dark-state conformations could describe the measured time-course SAXS data with a relaxation time τREC of ~ 34 to 35 min being larger than the recovery time found with UV/vis spectroscopy. Unlike the flavin chromophore-based UV/vis method that is sensitive to the local chromophore environment in flavoproteins, SAXS-based assay depends on protein conformational changes and provides with an alternative to measure the recovery kinetics.