scholarly journals Combining steady state and temperature jump IR spectroscopy to investigate the allosteric effects of ligand binding to dsDNA

2021 ◽  
Author(s):  
Jessica Dale ◽  
C. Peter Howe ◽  
Hedvika Toncrova ◽  
Robby Fritzsch ◽  
Gregory M Greetham ◽  
...  
Keyword(s):  
Steady State ◽  
Ir Spectroscopy ◽  
Ligand Binding ◽  
Experimental Evidence ◽  
Conformational Change ◽  
Structural Dynamics ◽  
Temperature Jump ◽  
Direct Experimental Evidence

Changes in the structural dynamics of double stranded (ds)DNA upon ligand binding have been linked to the mechanism of allostery without conformational change, but direct experimental evidence remains elusive. To...

Overexpression of cysteine dioxygenase reduces intracellular cysteine and glutathione pools in HepG2/C3A cells

2007 ◽  
Vol 293 (1) ◽  
pp. E62-E69 ◽  
Author(s):  
John E. Dominy ◽  
Jesse Hwang ◽  
Martha H. Stipanuk
Keyword(s):  
Steady State ◽  
Experimental Evidence ◽  
Cysteine Dioxygenase ◽  
Wild Type ◽  
Hepatic Expression ◽  
Direct Experimental Evidence ◽  
Homeostatic Response ◽  
Catabolic Enzyme ◽  
In Cells

Cysteine levels are carefully regulated in mammals to balance metabolic needs against the potential for cytotoxicity. It has been postulated that one of the major regulators of intracellular cysteine levels in mammals is cysteine dioxygenase (CDO). Hepatic expression of this catabolic enzyme increases dramatically in response to increased cysteine availability and may therefore be part of a homeostatic response to shunt excess toxic cysteine to more benign metabolites such as sulfate or taurine. Direct experimental evidence, however, is lacking to support the hypothesis that CDO is capable of altering steady-state intracellular cysteine levels. In this study, we expressed either the wild-type (WT) or a catalytically inactivated mutant (H86A) isoform of CDO in HepG2/C3A cells (which do not express endogenous CDO protein) and cultured them in different concentrations of extracellular cysteine. WT CDO, but not H86A CDO, was capable of reducing intracellular cysteine levels in cells incubated in physiologically relevant concentrations of cysteine. WT CDO also decreased the glutathione pool and potentiated the toxicity of CdCl2. These results demonstrate that CDO is capable of altering intracellular cysteine levels as well as glutathione levels.

scholarly journals Crystal structures of PI3K-C2α PX domain indicate conformational change associated with ligand binding

2008 ◽  
Vol 8 (1) ◽  
pp. 13 ◽  
Author(s):  
Gary N Parkinson ◽  
David Vines ◽  
Paul C Driscoll ◽  
Snezana Djordjevic
Keyword(s):  
Ligand Binding ◽  
Crystal Structures ◽  
Conformational Change ◽  
Px Domain

scholarly journals Experimental evidence of high pressure decoupling between charge transport and structural dynamics in a protic ionic glass-former

2017 ◽  
Vol 7 (1) ◽  
Author(s):  
Z. Wojnarowska ◽  
M. Rams-Baron ◽  
J. Knapik-Kowalczuk ◽  
A. Połatyńska ◽  
M. Pochylski ◽  
...  
Keyword(s):  
High Pressure ◽  
Charge Transport ◽  
Experimental Evidence ◽  
Structural Dynamics

scholarly journals Positive co-operative binding at two weak lysine-binding sites governs the Glu-plasminogen conformational change

1992 ◽  
Vol 285 (2) ◽  
pp. 419-425 ◽  
Author(s):  
U Christensen ◽  
L Mølgaard
Keyword(s):  
Ligand Binding ◽  
Binding Site ◽  
Conformational Change ◽  
Conformational Changes ◽  
Binding Sites ◽  
High Specificity ◽  
Stopped Flow ◽  
Protein Fluorescence ◽  
Lysine Residues ◽  
Kinetics Of

The kinetics of a series of Glu-plasminogen ligand-binding processes were investigated at pH 7.8 and 25 degrees C (in 0.1 M-NaCl). The ligands include compounds analogous to C-terminal lysine residues and to normal lysine residues. Changes of the Glu-plasminogen protein fluorescence were measured in a stopped-flow instrument as a function of time after rapid mixing of Glu-plasminogen and ligand at various concentrations. Large positive fluorescence changes (approximately 10%) accompany the ligand-induced conformational changes of Glu-plasminogen resulting from binding at weak lysine-binding sites. Detailed studies of the concentration-dependencies of the equilibrium signals and the rate constants of the process induced by various ligands showed the conformational change to involve two sites in a concerted positive co-operative process with three steps: (i) binding of a ligand at a very weak lysine-binding site that preferentially, but not exclusively, binds C-terminal-type lysine ligands, (ii) the rate-determining actual-conformational-change step and (iii) binding of one more lysine ligand at a second weak lysine-binding site that then binds the ligand more tightly. Further, totally independent initial small negative fluorescence changes (approximately 2-4%) corresponding to binding at the strong lysine-binding site of kringle 1 [Sottrup-Jensen, Claeys, Zajdel, Petersen & Magnusson (1978) Prog. Chem. Fibrinolysis Thrombolysis 3, 191-209] were observed for the C-terminal-type ligands. The finding that the conformational change in Glu-plasminogen involves two weak lysine-binding sites indicates that the effect cannot be assigned to any single kringle and that the problem of whether kringle 4 or kringle 5 is responsible for the process resolves itself. Probably kringle 4 and 5 are both participating. The involvement of two lysine binding-sites further makes the high specificity of Glu-plasminogen effectors more conceivable.

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