Site-Directed Mutagenesis of Tyrosine-98 in the Flavodoxin from Desulfovibrio vulgaris (Hildenborough): Regulation of Oxidation-Reduction Properties of the Bound FMN Cofactor by Aromatic, Solvent, and Electrostatic Interactions

Biochemistry ◽  
1994 ◽  
Vol 33 (28) ◽  
pp. 8505-8514 ◽  
Author(s):  
Richard P. Swenson ◽  
Grigorios D. Krey
Keyword(s):  
Electrostatic Interactions ◽  
Site Directed Mutagenesis ◽  
Desulfovibrio Vulgaris ◽  
Directed Mutagenesis ◽  
Aromatic Solvent ◽  
Oxidation Reduction ◽  
Desulfovibrio Vulgaris Hildenborough

scholarly journals Evidence for crucial electrostatic interactions between Bcl-2 homology domains BH3 and BH4 in the anti-apoptotic Nr-13 protein

2002 ◽  
Vol 368 (1) ◽  
pp. 213-221 ◽  
Author(s):  
Philippe LALLE ◽  
Abdel AOUACHERIA ◽  
Agnès DUMONT-MISCOPEIN ◽  
Martin JAMBON ◽  
Séverine VENET ◽  
...  
Keyword(s):  
Energy Minimization ◽  
Electrostatic Interactions ◽  
Family Members ◽  
Biological Significance ◽  
Terminal Region ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis ◽  
Conserved Domains ◽  
Apoptotic Activity ◽  
Interacting Domain

Nr-13 is an anti-apoptotic member of the Bcl-2 family previously shown to interact with Bax. The biological significance of this interaction was explored both in yeast and vertebrate cells and revealed that Nr-13 is able to counteract the pro-apoptotic activity of Bax. The Bax-interacting domain has been identified and corresponds to α-helices 5 and 6 in Nr-13. Site-directed mutagenesis has revealed that the N-terminal region of Nr-13 is essential for activity and corresponds to a genuine Bcl-2 homology domain (BH4). The modelling of Nr-13, based on its similarity with other Bcl-2 family proteins and energy minimization, suggests the possibility of electrostatic interactions between the two N-terminal-conserved domains BH4 and BH3. Disruption of these interactions severely affects Nr-13 anti-apoptotic activity. Together our results suggest that electrostatic interactions between BH4 and BH3 domains play a role in the control of activity of Nr-13 and a subset of Bcl-2 family members.

scholarly journals Significance of Local Electrostatic Interactions in Staphylococcal Nuclease Studied by Site-directed Mutagenesis

2001 ◽  
Vol 276 (49) ◽  
pp. 46039-46045 ◽  
Author(s):  
King Wong Leung ◽  
Yen-Chywan Liaw ◽  
Siu Chiu Chan ◽  
Hau Yi Lo ◽  
Faik N. Musayev ◽  
...  
Keyword(s):  
Electrostatic Interactions ◽  
Staphylococcal Nuclease ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis

scholarly journals Analysis of the ligand-binding domain of human retinoic acid receptor alpha by site-directed mutagenesis.

1996 ◽  
Vol 16 (10) ◽  
pp. 5386-5392 ◽  
Author(s):  
F P Lamour ◽  
P Lardelli ◽  
C M Apfel
Keyword(s):  
Retinoic Acid ◽  
Ligand Binding ◽  
Electrostatic Interactions ◽  
Cysteine Residue ◽  
Binding Domain ◽  
Ligand Binding Domain ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis ◽  
Arginine Residues ◽  
Antagonist Binding

Three subtypes of retinoic acid receptors (RAR), termed RAR alpha, RAR beta, and RAR gamma, have been described. They are composed of different structural domains, including distinct domains for DNA and ligand binding. RARs specifically bind all-trans-retinoic acid (RA), 9-cis-RA, and retinoid analogs. In this study, we examined the functional role of cysteine and arginine residues in the ligand-binding domain of hRAR alpha (hRAR alpha-LBD, amino acids 154 to 462). All conserved cysteine and arginine residues in this domain were mutated by site-directed mutagenesis, and the mutant proteins were characterized by blocking reactions, ligand-binding experiments, transactivation assays, and protease mapping. Changes of any cysteine residue of the hRAR alpha-LBD had no significant influence on the binding of all-trans RA or 9-cis RA. Interestingly, residue C-235 is specifically important in antagonist binding. With respect to arginine residues, only the two single mutations of R-276 and R-394 to alanine showed a dramatic decrease of agonist and antagonist binding whereas the R272A mutation showed only a slight effect. For all other arginine mutations, no differences in affinity were detectable. The two mutations R217A and R294A caused an increased binding efficiency for antagonists but no change in agonist binding. From these results, we can conclude that electrostatic interactions of retinoids with the RAR alpha-LBD play a significant role in ligand binding. In addition, antagonists show distinctly different requirements for efficient binding, which may contribute to their interference in the ligand-inducible transactivation function of RAR alpha.

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