Differential scanning calorimetry as a tool to estimate binding parameters in multiligand binding proteins

Celej, M. Soledad; Dassie, Sergio Alberto; González, Martı́n; Bianconi, M. Lucia; Fidelio, Gerardo D.

doi:10.1016/j.ab.2005.12.029

Cited by 64 publications

(42 citation statements)

References 26 publications

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“…27) They demonstrated that thermal stability of HSA monomer, monitored by DSC, decreased with increasing protein concentration. 27) However, mean T m value of 73.7°C for all HSA products in this study was higher than those of the previous studies described above (59.2-63.1°C) 15,25,26) because medical-grade HSA products typically contain the albumin-specific stabilizers N-Ac Trp and caprylate, in order to confer resistance to pasteurization (60°C for 10 h) for viral inactivation. 6) In other words, the addition of stabilizers may be of major importance due to their effects on the thermal stability of proteins, and medicalgrade HSA products have thermally more stable structures than HSA with no additives (native state).…”

Section: Discussioncontrasting

confidence: 54%

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Quantitation of Oxidative Modifications of Commercial Human Albumin for Clinical Use

Takahashi

Terada

Arikawa

et al. 2016

Biological & Pharmaceutical Bulletin

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We investigated the quantitation of oxidative chemical modifications, such as thiol oxidation and carbonylation, in medical-grade human serum albumin (HSA) preparations, in comparison with those of healthy and diseased subjects. Four kinds of HSA products were obtained from three major suppliers in Japan. Eight male collegiate students and six healthy male volunteers were recruited as the young (21.6 years) and older (57.2 years) groups, respectively. Four male stable patients (64.3 years) treated with regular hemodialysis (HD) also enrolled in this study. Quantitative analyses for thiol oxidation and carbonylation were performed using HPLC and spectroscopic methods, respectively. Structural characterization was further investigated by differential scanning calorimetry (DSC) and circular dichroism (CD) spectropolarimetry. Significantly larger amounts of thiol-oxidized and carbonylated HSA products were observed than HSA obtained from healthy subjects. In the structural characterization, the midpoint temperature of the denaturation curve (T m ) analyzed by DSC was relatively high, and may have been caused by the added albumin-specific stabilizers, and CD-resolved secondary structure showed that HSA products had a helical conformation. Commercial HSA products for clinical use have a more thermally stable state and remain in a helix-rich structure, even though their specific amino acids (mainly Cys and Lys residues) are oxidatively modified.

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Section: Discussioncontrasting

confidence: 54%

“…Co.). 26) DSC thermogram for the HSA products in this study showed a monophasic endotherm (data not shown), and the T m values for HSA products ranged between 72 and 75°C, as shown in Table 2. Among them, the T m value for product (#3), i.e., 'Benesis-25%,' was significantly lower than those for the other three products (p<0.001).…”

Section: Discussionmentioning

confidence: 97%

Quantitation of Oxidative Modifications of Commercial Human Albumin for Clinical Use

Takahashi

Terada

Arikawa

et al. 2016

Biological & Pharmaceutical Bulletin

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“…This sequence was coupled to a multidimensional optimization routine in order to get K b app (see ref. 28 and references therein for details). The intrinsic unfolding parameters were obtained from an endotherm of free BSA.…”

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confidence: 99%

“…We have successfully used this approach to estimate the number of binding sites for different dyes on serum albumins. 24,28) In order to confirm the preferential binding of AHTC to native BSA, as suggested by our calorimetric results (Fig. 5, Table 2), the emission spectra of free and bound AHTC at different temperatures were compared.…”

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confidence: 99%

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Binding of the Highly Toxic Tetracycline Derivative, Anhydrotetracycline, to Bovine Serum Albumin

Burgos

Fernández

Celej

et al. 2011

Biological & Pharmaceutical Bulletin

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Tetracycline (TC) derivatives comprise a group of bacteriostatic antibiotics extensively used in human and veterinary medicine and in acquaculture, and as animal growth promoters due to their safe toxicological profile.1) More recently, they have been proposed as anti-amyloidogenic drugs since they can interfere with protein conformational changes and self-assemblies associated with protein misfolding and deposition diseases. 2-4)Despite their popular use, TC derivatives are highly unstable and some degradation products formed during the storage or normal use of the drugs are pharmacologically toxic or inactive compounds.5) Anhydrotetracycline (AHTC) is one of the major degradation products of TC that has been linked to several side effects of the drug, e.g., cutaneous phototoxicity 6) and Fanconi-type syndrome. 7) AHTC has also proven to be more toxic than TC against environmentally relevant bacteria. 8)AHTC can be produced by dehydration in acid media, 5,8) by photolysis in mild reducing conditions 6,9,10) or during high-temperature treatments.11) Removal of a water molecule from the C5a-C6 positions of TC by treatment with warm mineral acid gives the anhydroderivative (Fig. 1).12) Dehydration leads to significant changes in aromatization and planarity of the tetracyclic moiety, 13,14) showing variations in potency, 15) mechanism of action 1) and lipophilicity 16,17) as compared to the parent compound. Changes in chemical configuration can also modulate the interaction of drugs with serum proteins. 18)Serum albumins are the main carriers of physiological metabolites and pharmacological drugs in blood. Protein binding has a significant impact on the pharmacokinetics and biological activity of drugs and modulates the toxicity of bound substances, attenuates adverse reactions and prolongs the in vivo half-life of therapeutic agents. 18-21)The facts outlined above prompted us to investigate the interaction of AHTC with bovine serum albumin (BSA) using three biophysical techniques. Binding affinity and number of interacting ligands were assessed by fluorimetric titration. . Drug binding was enthalpically and entropically driven and seemed to involve hydrophobic interactions. AHTC fluorescence enhancement and hypsochromic shifts observed upon binding suggested a low-polarity location excluded from water for the bound drug. Our data are useful for evaluating the biodisponibility of the pharmacophore and the dynamic distribution of the toxic derivative.

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Thermodynamic and biophysical study of fatty acid effector binding to soybean lipoxygenase: implications for allostery driven by helix α2 dynamics

et al. 2022

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Previous comparative kinetic isotope effects have inferred an allosteric site for fatty acids and their derivatives that modulates substrate selectivity in 15lipoxygenases. Hydrogen-deuterium exchange also previously revealed regionally defined enhanced protein flexibility, centred at helix a2a gate to the substrate entrance. Direct evidence for allosteric binding and a complete understanding of its mechanism remains elusive. In this study, we examine the binding thermodynamics of the fatty acid mimic, oleyl sulfate (OS), with the monomeric model plant 15-LOX, soybean lipoxygenase (SLO), using isothermal titration calorimetry. Dynamic light scattering and differential scanning calorimetry rule out OS-induced oligomerization or structural changes. These data provide evidence that the fatty acid allosteric regulation of SLO is controlled by the dynamics of helix a2.

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Differential scanning calorimetry as a tool to estimate binding parameters in multiligand binding proteins

Cited by 64 publications

References 26 publications

Quantitation of Oxidative Modifications of Commercial Human Albumin for Clinical Use

Quantitation of Oxidative Modifications of Commercial Human Albumin for Clinical Use

Binding of the Highly Toxic Tetracycline Derivative, Anhydrotetracycline, to Bovine Serum Albumin

Thermodynamic and biophysical study of fatty acid effector binding to soybean lipoxygenase: implications for allostery driven by helix α2 dynamics

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