scholarly journals Adenosine nucleotide binding sites on Complex V from diverse organisms

2021 ◽  
Author(s):  
Abhinav Parashar ◽  
Kelath Murali Manoj
Keyword(s):  
Binding Sites ◽  
In Silico ◽  
Nucleotide Binding ◽  
Atp Binding ◽  
Atp Binding Site ◽  
In Silico Docking ◽  
Epsilon Subunit ◽  
Nucleotide Binding Sites ◽  
Alpha Beta ◽  
Adenosine Nucleotide

Using in silico docking approaches, we scan the various subunits of Complex V (FoF1ATPase) for putative adenosine nucleotide binding sites. We find that multiple generic ADP/ATP binding sites are present on the alpha-beta binding sites and a conserved ATP binding site is present on the epsilon subunit. These findings support the murburn model of Complex V.

scholarly journals The properties of Msh2–Msh6 ATP binding mutants suggest a signal amplification mechanism in DNA mismatch repair

2018 ◽  
Vol 293 (47) ◽  
pp. 18055-18070 ◽  
Author(s):  
William J. Graham ◽  
Christopher D. Putnam ◽  
Richard D. Kolodner
Keyword(s):  
Mismatch Repair ◽  
Binding Sites ◽  
Signal Amplification ◽  
Dna Mismatch Repair ◽  
Nucleotide Binding ◽  
Atp Binding ◽  
Dna Mismatch ◽  
Nucleotide Binding Sites ◽  
Sliding Clamp ◽  
Atp Concentration

DNA mismatch repair (MMR) corrects mispaired DNA bases and small insertion/deletion loops generated by DNA replication errors. After binding a mispair, the eukaryotic mispair recognition complex Msh2–Msh6 binds ATP in both of its nucleotide-binding sites, which induces a conformational change resulting in the formation of an Msh2–Msh6 sliding clamp that releases from the mispair and slides freely along the DNA. However, the roles that Msh2–Msh6 sliding clamps play in MMR remain poorly understood. Here, using Saccharomyces cerevisiae, we created Msh2 and Msh6 Walker A nucleotide–binding site mutants that have defects in ATP binding in one or both nucleotide-binding sites of the Msh2–Msh6 heterodimer. We found that these mutations cause a complete MMR defect in vivo. The mutant Msh2–Msh6 complexes exhibited normal mispair recognition and were proficient at recruiting the MMR endonuclease Mlh1–Pms1 to mispaired DNA. At physiological (2.5 mm) ATP concentration, the mutant complexes displayed modest partial defects in supporting MMR in reconstituted Mlh1–Pms1-independent and Mlh1–Pms1-dependent MMR reactions in vitro and in activation of the Mlh1–Pms1 endonuclease and showed a more severe defect at low (0.1 mm) ATP concentration. In contrast, five of the mutants were completely defective and one was mostly defective for sliding clamp formation at high and low ATP concentrations. These findings suggest that mispair-dependent sliding clamp formation triggers binding of additional Msh2–Msh6 complexes and that further recruitment of additional downstream MMR proteins is required for signal amplification of mispair binding during MMR.

Discrete mutations introduced in the predicted nucleotide-binding sites of the mdr1 gene abolish its ability to confer multidrug resistance

1989 ◽  
Vol 9 (12) ◽  
pp. 5289-5297
Author(s):  
M Azzaria ◽  
E Schurr ◽  
P Gros
Keyword(s):  
Drug Resistance ◽  
Biological Activity ◽  
Multidrug Resistance ◽  
Amino Acid ◽  
Binding Sites ◽  
Consensus Sequence ◽  
Nucleotide Binding ◽  
Atp Binding ◽  
Nucleotide Binding Sites ◽  
The Core

In cells stably transfected and overexpressing the mouse mdr1 gene, multidrug resistance is associated with an increased ATP-dependent drug efflux. Analysis of the predicted amino acid sequence of the MDR1 protein revealed the presence of two putative nucleotide-binding sites (NBS). To assess the functional importance of these NBS in the overall drug resistance phenotype conferred by mdr1, we introduced amino acid substitutions in the core consensus sequence for nucleotide binding, GXGKST. Mutants bearing the sequence GXAKST or GXGRST at either of the two NBS of mdr1 and a double mutant harboring the sequence GXGRST at both NBS were generated. The integrity of the two NBS was essential for the biological activity of mdr1, since all five mutants were unable to confer drug resistance to hamster drug-sensitive cells in transfection experiments. Conversely, a lysine-to-arginine substitution outside the core consensus sequence had no effect on the activity of mdr1. Failure to reduce intracellular accumulation of [3H]vinblastine paralleled the loss of activity in cell clones expressing mutant MDR1 proteins. However, the ability to bind the photoactivatable ATP analog 8-azido ATP was retained in the five inactive MDR1 mutants. This result implies that an essential step subsequent to ATP binding is impaired in these mutants, possibly ATP hydrolysis or secondary conformational changes induced by ATP-binding or hydrolysis. Our results suggest that the two NBS function in a cooperative fashion, since mutations in a single NBS completely abrogated the biological activity of mdr1.

scholarly journals Dependence on Membrane Lipids of the Effect of Vanadate on Calcium and ATP Binding to Sarcoplasmic Reticulum ATPase

1984 ◽  
Vol 39 (11-12) ◽  
pp. 1137-1140 ◽  
Author(s):  
Pankaj Medda ◽  
Wilhelm Hasselbach
Keyword(s):  
Sarcoplasmic Reticulum ◽  
Binding Sites ◽  
Calcium Binding ◽  
Membrane Lipids ◽  
Nucleotide Binding ◽  
Atp Binding ◽  
Transport Atpase ◽  
Nucleotide Binding Sites ◽  
High Affinity ◽  
Calcium Binding Sites

Abstract The affinity of the sarcoplasmic reticulum transport ATPase for calcium and ATP is not affected by lipid depriviation while vanadate binding is completely abolished. Lipid substitution restores vanadate binding as well as the vanadate induced disappearance of the enzyme’s high affinity calcium and nucleotide binding sites. Nucleotide binding is simultaneously restored with the displacement of vanadate from the enzyme following the occupation of its low affinity calcium binding sites.

scholarly journals Discrete mutations introduced in the predicted nucleotide-binding sites of the mdr1 gene abolish its ability to confer multidrug resistance.

1989 ◽  
Vol 9 (12) ◽  
pp. 5289-5297 ◽  
Author(s):  
M Azzaria ◽  
E Schurr ◽  
P Gros
Keyword(s):  
Drug Resistance ◽  
Biological Activity ◽  
Multidrug Resistance ◽  
Amino Acid ◽  
Binding Sites ◽  
Consensus Sequence ◽  
Nucleotide Binding ◽  
Atp Binding ◽  
Nucleotide Binding Sites ◽  
The Core

In cells stably transfected and overexpressing the mouse mdr1 gene, multidrug resistance is associated with an increased ATP-dependent drug efflux. Analysis of the predicted amino acid sequence of the MDR1 protein revealed the presence of two putative nucleotide-binding sites (NBS). To assess the functional importance of these NBS in the overall drug resistance phenotype conferred by mdr1, we introduced amino acid substitutions in the core consensus sequence for nucleotide binding, GXGKST. Mutants bearing the sequence GXAKST or GXGRST at either of the two NBS of mdr1 and a double mutant harboring the sequence GXGRST at both NBS were generated. The integrity of the two NBS was essential for the biological activity of mdr1, since all five mutants were unable to confer drug resistance to hamster drug-sensitive cells in transfection experiments. Conversely, a lysine-to-arginine substitution outside the core consensus sequence had no effect on the activity of mdr1. Failure to reduce intracellular accumulation of [3H]vinblastine paralleled the loss of activity in cell clones expressing mutant MDR1 proteins. However, the ability to bind the photoactivatable ATP analog 8-azido ATP was retained in the five inactive MDR1 mutants. This result implies that an essential step subsequent to ATP binding is impaired in these mutants, possibly ATP hydrolysis or secondary conformational changes induced by ATP-binding or hydrolysis. Our results suggest that the two NBS function in a cooperative fashion, since mutations in a single NBS completely abrogated the biological activity of mdr1.

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