Nano-second protein dynamics of key residue at Position 38 in catechol-O-methyltransferase system: a time-resolved fluorescence study

2020 ◽  
Vol 168 (4) ◽  
pp. 417-425
Author(s):  
Fan Liu ◽  
Jianyu Zhang
Keyword(s):  
Protein Dynamics ◽  
Enzyme Catalysis ◽  
Time Resolved Fluorescence ◽  
Fluorescence Study ◽  
Time Resolved ◽  
System A ◽  
Key Enzyme ◽  
Relationship Of ◽  
The Relationship ◽  
Stokes Shifts

Abstract Human catechol-O-methyltransferase, a key enzyme related to neurotransmitter metabolism, catalyses a methyl transfer from S-adenosylmethionine to catechol. Although extensive studies aim to understand the enzyme mechanisms, the connection of protein dynamics and enzyme catalysis is still not clear. Here, W38in (Trp143Phe) and W38in/Y68A (Trp143Phe with Tyr68Ala) mutants were carried out to study the relationship of dynamics and catalysis in nano-second timescale using time-resolved fluorescence lifetimes and Stokes shifts in various solvents. The comprehensive data implied the mutant W38in/Y68A with lower activity is more rigid than the ‘WT’−W38in, suggesting the importance of flexibility at residue 38 to maintain the optimal catalysis.

The time-resolved fluorescence study of kinetics and thermodynamics of Eu(iii) and Tb(iii) complexes with the DO2A macrocyclic ligand

2018 ◽  
Vol 42 (10) ◽  
pp. 7993-8000
Author(s):  
Filip Smrčka ◽  
Přemysl Lubal
Keyword(s):  
Fluorescence Spectroscopy ◽  
Macrocyclic Ligand ◽  
Time Resolved Fluorescence ◽  
Fluorescence Study ◽  
Time Resolved ◽  
Kinetics And Thermodynamics ◽  
Thermodynamics And Kinetics ◽  
Kinetics Of Formation ◽  
Kinetics Of

The thermodynamics and kinetics of formation/dissociation of Eu(iii) and Tb(iii) with the H2DO2A macrocyclic ligand were studied by time-resolved fluorescence spectroscopy.

Tryptophan interactions of gramicidin A′ channels in lipids: a time-resolved fluorescence study

1986 ◽  
Vol 862 (2) ◽  
pp. 265-272 ◽  
Author(s):  
Lanfranco Masotti ◽  
Paolo Cavatorta ◽  
Giorgio Sartor ◽  
Emanuela Casali ◽  
Arthur G. Szabo
Keyword(s):  
Gramicidin A ◽  
Time Resolved Fluorescence ◽  
Fluorescence Study ◽  
Time Resolved

scholarly journals High-Throughput Screen for Inhibitors of 1-Deoxy-D-Xylulose 5-Phosphate Reductoisomerase by Surrogate Ligand Competition

2003 ◽  
Vol 8 (3) ◽  
pp. 332-339 ◽  
Author(s):  
Elizabeth B. Gottlin ◽  
R. Edward Benson ◽  
Scott Conary ◽  
Brett Antonio ◽  
Kellie Duke ◽  
...  
Keyword(s):  
High Throughput ◽  
Binding Sites ◽  
Biosynthetic Pathway ◽  
Enzyme Assay ◽  
High Throughput Screen ◽  
Time Resolved Fluorescence ◽  
Time Resolved ◽  
Biochemical Assays ◽  
Key Enzyme ◽  
Nadph Oxidation

1-Deoxy-D-xylulose 5-phosphate reductoisomerase (Dxr) is a key enzyme in a biosynthetic pathway for isoprenoids that is unique to eubacteria and plants. Dxr catalyzes the rearrangement and NADPH-dependent reduction of 1-deoxy-D-xylulose 5-phosphate to 2-C-methyl-D-erythritol 4-phosphate. The authors have purified Escherichia coli Dxr and devised a high-throughput screen (HTS) for compounds that bind to this enzyme at a functional site. Evidence is presented that the surrogate ligand directly binds or allosterically affects both the D-1-deoxyxylulose 5-phosphate (DXP) and NADPH binding sites. Compounds that bind at either or both sites that compete for binding with the surrogate ligand register as hits. The time-resolved fluorescence-based assay represents an improvement over the Dxr enzyme assay that relies on relatively insensitive measurements of NADPH oxidation. Screening 32,000 compounds from a diverse historical library, the authors obtained 89 potent inhibitors in the surrogate ligand competition assay. The results presented here suggest that peptide surrogate ligands may be useful in formatting HTS for proteins with difficult biochemical assays or targets of unknown function. ( Journal of Biomolecular Screening 2003:332-339)

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