title: Temperature dependence of ligand-protein complex formation as reflected by saturation transfer difference NMR experiments.
abstract: We show that temperature is an important parameter for the sensitivity of saturation transfer difference (STD) spectroscopy. A decreased intensity of STD signals is observed for lactose binding to growth-regulatory galectin7 (p53-induced gene 1), as well as for nucleotide binding to annexin A6, when the temperature is increased from 281 to 298-310 K. Opposite temperature effects on STD intensity are observed for S-peptide binding to S-protein to reconstitute RNase S. However, the STD signals for tryptophan binding to downstream regulatory element antagonist modulator of the human prodynorphin gene (DREAM)are relatively unaffected between 281 and 298 K. The known kinetics of the binding of ATP by the uncoupling protein from brown adipose tissue mitochondria (UCP1) predicted an observable STD at 310 K, but rapid sample degradation limits the experiments to much lower temperatures. Temperature strongly influences the kinetics and affinity constant of various types of complex formation and in so doing influences the observed STD effects. Therefore, temperature can be exploited to facilitate the optimization of STD-based applications, and at the same time minimize the number of test samples. STD-based screening protocols to detect new target-specific compounds may yield a larger number of potential ligands if screened at various temperatures.
authors.affiliation: Department of Protein Science, Centro de Investigaciones Biológicas, CSIC, Ramiro de Maeztu 9, 28040 Madrid, Spain. p.groves@ulster.ac.uk
authors.forename: Patrick
authors.initials: P
authors.lastname: Groves
authors.forename: Katalin E
authors.initials: KE
authors.lastname: Kövér
authors.forename: Sabine
authors.initials: S
authors.lastname: André
authors.forename: Joanna
authors.initials: J
authors.lastname: Bandorowicz-Pikula
authors.forename: Gyula
authors.initials: G
authors.lastname: Batta
authors.forename: Marta
authors.initials: M
authors.lastname: Bruix
authors.forename: René
authors.initials: R
authors.lastname: Buchet
authors.forename: Angeles
authors.initials: A
authors.lastname: Canales
authors.forename: F Javier
authors.initials: FJ
authors.lastname: Cañada
authors.forename: Hans-Joachim
authors.initials: HJ
authors.lastname: Gabius
authors.forename: Douglas V
authors.initials: DV
authors.lastname: Laurents
authors.forename: Jose R
authors.initials: JR
authors.lastname: Naranjo
authors.forename: Małgorzata
authors.initials: M
authors.lastname: Palczewska
authors.forename: Slawomir
authors.initials: S
authors.lastname: Pikula
authors.forename: Eduardo
authors.initials: E
authors.lastname: Rial
authors.forename: Agnieszka
authors.initials: A
authors.lastname: Strzelecka-Kiliszek
authors.forename: Jesús
authors.initials: J
authors.lastname: Jiménez-Barbero
mesh_terms: D017317:Annexin A6 / Q000737:chemistry; D007473:Ion Channels / Q000737:chemistry; D037102:Lectins / Q000737:chemistry; D008024:Ligands; D009682:Magnetic Resonance Spectroscopy; D024101:Mitochondrial Proteins / Q000737:chemistry; D011506:Proteins / Q000737:chemistry*; D011522:Protons; D013696:Temperature; D013816:Thermodynamics; D000071256:Uncoupling Protein 1
chemical_list: D017317:Annexin A6; D007473:Ion Channels; D037102:Lectins; D008024:Ligands; D024101:Mitochondrial Proteins; D011506:Proteins; D011522:Protons; C000604371:UCP1 protein, human; D000071256:Uncoupling Protein 1