Stability studies on pig heart mitochondrial malate dehydrogenase: the effect of salts and amino acids

Jensen, Wayne A.; Armstrong, J. M.; Giorgio, Joseph; Hearn, Milton Thomas William

doi:10.1016/0167-4838(96)00049-0

Cited by 33 publications

(16 citation statements)

References 51 publications

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“…The proteins involved in above studies were bovine serum albumin (10)(11), L-malate dehydrogenase (MDH) (11), and pig heart mitochondrial malate dehydrogenase (phm-MDH) (12,13). These studies indicate that both the cations and anions play an important role in protein stabilization or its inactivation.…”

Section: Introductionmentioning

confidence: 94%

Model for Inactivation of α‐Amylase in the Presence of Salts: Theoretical and Experimental Studies

Lecker

Khan

1998

Biotechnology Progress

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According to a previous report, only the smaller anions, like chlorides,that readily fit into the anion binding site of alpha-amylase can cause an increased stability (relative to enzymes in aqueous solution), and the anions that are too large to fit into the binding site should have no effect on the enzyme. Even though the results on large benzoate ions are consistent with the above postulate, much larger citrate ions from sodium and potassium citrate show stabilization at moderate salt concentrations and follow an expected trend of low stability only at large salt concentrations. The citrate ions from ammonium citrate exhibit very little to almost no stabilization. In addition, low to moderate concentrations of NaCl that provide a large stability to the enzyme show almost no stability in the presence of EDTA. We put forward an inactivation model that involves a reversible dissociation of the anion bound to the protein, followed by a reversible inactivation step of calcium ion dissociation and an irreversible denaturation step of apoenzyme.

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Section: Introductionmentioning

confidence: 94%

Model for Inactivation of α‐Amylase in the Presence of Salts: Theoretical and Experimental Studies

Lecker

Khan

1998

Biotechnology Progress

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“…Among carboxylic acid salts, acetate and citrate have long been known to act as protein stabilizers. In some studies,4–11 the diversification of citrate as stabilizer for several biomolecules can be observed.…”

Section: Introductionmentioning

confidence: 99%

Citrate and phosphate influence on green fluorescent protein thermal stability

Novaes

Mazzola

Pessoa

et al. 2010

Biotechnology Progress

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Protein structure and function can be regulated by no specific interactions, such as ionic interactions in the presence of salts. Green fluorescent protein (GFP) shows remarkable structural stability and high fluorescence; its stability can be directly related to its fluorescence output, among other characteristics. GFP is stable under increasing temperatures, and its thermal denaturation is highly reproducible. The aim of this study was to evaluate the thermal stability of GFP in the presence of different salts at several concentrations and exposed to constant temperatures, in a range of 70-95°C. Thermal stability was expressed in decimal reduction time. It was observed that the D-values obtained were higher in the presence of citrate and phosphate, when compared with that obtained in their absence, indicating that these salts stabilized the protein against thermal denaturation.

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“…31 MDH is a dimeric protein with 8 cysteine residues per 34 kDa subunit and has no disulfide bonds. 32 Immunoblotting demonstrated that GRd and MDH were modified by NB-DCVCS. These results suggest that it is possible to carry out studies involving the detection of covalent modification of specific proteins by NB-DCVCS in vitro.…”

Section: Discussionmentioning

confidence: 99%

N-Biotinyl-S-(1,2-dichlorovinyl)-l-cysteine Sulfoxide as a Potential Model for S-(1,2-Dichlorovinyl)-l-cysteine Sulfoxide: Characterization of Stability and Reactivity with Glutathione and Kidney Proteins in Vitro

Irving

Brownfield²,

Elfarra³

2011

Chem. Res. Toxicol.

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S-(1,2-Dichlorovinyl)-L-cysteine sulfoxide (DCVCS) is a reactive and potent nephrotoxic metabolite of the human trichloroethylene metabolite S-(1,2-dichlorovinyl)-L-cysteine (DCVC). Because DCVCS covalent binding to kidney proteins likely plays a role in its nephrotoxicity, in this study biotin-tagged DCVCS, N-Biotinyl-DCVCS (NB-DCVCS), was synthesized and its stability in buffer alone and in the presence of rat blood or plasma was characterized in vitro. In addition, reactivity toward GSH and covalent binding to selected model enzymes and isolated kidney proteins were characterized. The half-lives of NB-DCVCS (39.6 min) and the DCVCS (diastereomer 1: 14.4 min, diastereomer 2: 6 min) in the presence of GSH were comparable. Incubating the model enzymes glutathione reductase and malate dehydrogenase with 10 µM NB-DCVCS for 3 h at 37°C followed by immunoblotting using anti-biotin antibodies demonstrated that glutathione reductase and malate dehydrogenase were extensively modified by NB-DCVCS. When rat kidney cytosol (6 µg/µL) was incubated with NB-DCVCS (312.5 nM to 5 µM) for 3 h at 37°C followed by immunoblotting, a concentration-dependent increase in signal with multiple proteins with different molecular weights was observed, suggesting NB-DCVCS binds to multiple kidney proteins with different selectivity. Incubating rat kidney cytosol with DCVCS (10 – 100 µM) prior to addition of NB-DCVCS (2.5 µM) reduced the immunoblotting signal, suggesting that NB-DCVCS and DCVCS compete for the same binding sites. A comparison of the stability of NB-DCVCS and DCVCS in rat blood and plasma was determined in vitro and NB-DCVCS exhibited higher stability than DCVCS in both media. Collectively, these results suggest NB-DCVCS shows sufficient stability, reactivity, and selectivity to warrant further investigations into its possible use as a tool for future characterization of the role of covalent modification of renal proteins by DCVCS in nephrotoxicity.

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Stability studies on pig heart mitochondrial malate dehydrogenase: the effect of salts and amino acids

Cited by 33 publications

References 51 publications

Model for Inactivation of α‐Amylase in the Presence of Salts: Theoretical and Experimental Studies

Model for Inactivation of α‐Amylase in the Presence of Salts: Theoretical and Experimental Studies

Citrate and phosphate influence on green fluorescent protein thermal stability

N-Biotinyl-S-(1,2-dichlorovinyl)-l-cysteine Sulfoxide as a Potential Model for S-(1,2-Dichlorovinyl)-l-cysteine Sulfoxide: Characterization of Stability and Reactivity with Glutathione and Kidney Proteins in Vitro

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