Proposed temperature-dependent conformational transition in D-amino acid oxidase: a differential scanning microcalorimetric study.

Sturtevant, Julian M.; Mateo, Pedro L.

doi:10.1073/pnas.75.6.2584

Cited by 25 publications

(13 citation statements)

References 11 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…How can a substantial enthalpy change be detected by this spectroscopic analysis and not be calorimetry? Similar behavior has been observed for the protein d-amino-oxidase by Sturtevant and Mateo (1978) where the authors verified a reported fluorometric AH for the protein transition (Massey et al, 1965) but could not detect an enthalpy change calorimetrically. The discrepancies between the spectroscopic and calorimetric enthalpies can only mean that the microscopic AH as seen by changes in fluorophore environment does not reflect the true enthalpy change but rather relates to cooperative unit of the transition (Edsall and Gutfreund, 1983).…”

Section: Discussionsupporting

confidence: 79%

Evaluation of the thermal coefficient of the resistance to fluorophore rotation in model membranes

Scarlata

1989

Biophysical Journal

View full text Add to dashboard Cite

The thermal coefficient of the frictional resistance to fluorophore rotation (b), a parameter related to the change in the local viscosity with temperature, was determined for anthroyloxy-fatty acid probes in micelles and dimyristoyl lecithin (DMPC) and dioleoyl lecithin (DOPC) unilamellar and multilamellar vesicles. The value of b and the percent change in anisotropy with temperature (%dA/dT) remained constant with membrane depth and only depended on composition. These parameters were also the same when either in-plane, or in-plane and out-of-plane fluorophore motions were observed. This result indicates that the membranes expand isotropically. The magnitude of b was found to be primarily dependent on the packing of the hydrocarbon chains with higher b values relating to more closely-packed chains. b was responsive to the gel to liquid crystal phase transition of DMPC and the bilayer to hexagonal phase transition of egg-phosphatidylethanolamine. When the enthalpy values for the fluorophore transfer from one phase to another are calculated, the values are larger than those measured by calorimetry and reflect a discrepancy between the microscopic enthalpy experienced by the fluorophore due to a change in environment versus the macroscopic enthalpy of the system as a whole.

show abstract

Section: Discussionsupporting

confidence: 79%

Evaluation of the thermal coefficient of the resistance to fluorophore rotation in model membranes

Scarlata

1989

Biophysical Journal

View full text Add to dashboard Cite

show abstract

“…The binding interaction between apoproteins and flavin prosthetic groups has been studied extensively. The strong and specific binding of FMN or FAD to apoflavoproteins is driven by the enthalpic contribution to the free energy change of binding [40–42]. The thermodynamics of flavin binding to flavodoxin have been studied in detail [42–46].…”

Section: Thermodynamics Of Flavin Bindingmentioning

confidence: 99%

Deflavination and reconstitution of flavoproteins

Hefti¹,

Vervoort²,

Berkel³

2003

European Journal of Biochemistry

118

104

View full text Add to dashboard Cite

Flavoproteins are ubiquitous redox proteins that are involved in many biological processes. In the majority of flavoproteins, the flavin cofactor is tightly but noncovalently bound. Reversible dissociation of flavoproteins into apoprotein and flavin prosthetic group yields valuable insights in flavoprotein folding, function and mechanism. Replacement of the natural cofactor with artificial flavins has proved to be especially useful for the determination of the solvent accessibility, polarity, reaction stereochemistry and dynamic behaviour of flavoprotein active sites. In this review we summarize the advances made in the field of flavoprotein deflavination and reconstitution. Several sophisticated chromatographic procedures to either deflavinate or reconstitute the flavoprotein on a large scale are discussed. In a subset of flavoproteins, the flavin cofactor is covalently attached to the polypeptide chain. Studies from riboflavindeficient expression systems and site-directed mutagenesis suggest that the flavinylation reaction is a post-translational, rather than a cotranslational, process. These genetic approaches have also provided insight into the mechanism of covalent flavinylation and the rationale for this atypical protein modification.

show abstract

“…Their calorimetric studies, however, failed to show any concomitant change in heat capacity or enthalpy ofthe protein under identical conditions. This unequivocally rules out any thermodynamic transition and suggests that the fluorescence intensity reflects something other than the average conformational state of the protein (2).…”

mentioning

confidence: 99%

“…Given a fluorescence change, and assuming it to be a result of an equilibrium change in conformation, one might obtain the equilibrium constant for the transition from: K=F0 F(T) 2 F(7) [2] in which FO and F,,. are the low-and high-temperature limits of the fluorescence, respectively.…”

mentioning

confidence: 99%

“…are the low-and high-temperature limits of the fluorescence, respectively. The equilibrium constant would be related to the thermodynamic parameters for the transition in the usual way: K = exp(-AG/RT) = exp(AS/R)exp(-AH/RT), and a linear van Sturtevant (2) and by Londesborough (16). It now appears that even the apparently more direct probes of protein conformation will have to be examined more circumspectly, and similar problems may afflict techniques that purport to measure lipid phase transitions in biological membranes.…”

mentioning

confidence: 99%

See 1 more Smart Citation

Spurious conformational transitions in proteins?

Cooper¹

1981

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

Temperature-dependent dynamic processes in biological macromolecules can produce sharp and reversible transitions in spectroscopic properties that might be misinterpreted as evidence for thermally induced conformational changes. This provides a rational explanation for the paradoxical case ofD-amino acid oxidase [D-amino-acid (1) found a 30% decrease in tryptophan fluorescence intensity upon raising the temperature ofthe protein solution from 80C to 20TC. The transition was sharp and reversible, with a midpoint at about 14WC, and was interpreted as a thermally induced isomerization between two distinct conformational states of the protein separated by an enthalpy difference, AH, of about 78 kcal mold (1 kcal = 4.18 kJ). Interpretation ofthe more ambiguous discontinuities in absorbance and kinetic properties of the enzyme in the same temperature region (1) was complicated by subunit and cofactor dissociation equilibria. Subsequent work by Sturtevant and Mateo (2) confirmed the fluorescence transition, though with a lower apparent AH ofabout 50 kcal mol' for the flavoprotein, and 32 kcal mol-1 for the apoenzyme. Their calorimetric studies, however, failed to show any concomitant change in heat capacity or enthalpy ofthe protein under identical conditions. This unequivocally rules out any thermodynamic transition and suggests that the fluorescence intensity reflects something other than the average conformational state of the protein (2).Fluorescence is a stochastic phenomenon. It depends not only on intrinsic properties of the fluorophore but also on dynamic rate processes involving transient events such as molecular collisions, reorientations, and energy transfer during the lifetime ofthe fluorescent excited state (3-7). Various functional groups within proteins are capable ofquenching tryptophan fluorescence, including amines, carboxylic acids, sulfhydryls, and imidazole groups. Additional quenching can arise from energy transfer between suitably oriented aromatic groups and by interaction with solvent and solute molecules (3-7). All these can be dynamic processes requiring only transient interaction with the excited group. They may, therefore, be temperature dependent.The fluorescence quantum yield, F, of a species A may be written:F= kf kf + knr' in which kf and knr are the rate constants for the radiative and nonradiative decay processes (h is the Planck constant): kf A* ----*A + hp A* ---+ A + heat. Nonradiative decay may occur by a variety of mechanisms. Intrinsic processes, including intersystem crossing and internal conversion, will depend on the chemical nature ofthe molecule and bulk properties ofthe environment (polarity, dielectric constant, etc.) and will be relatively insensitive to changes in temperature per se, though they might be affected indirectly by conformational changes in a protein that modify the surroundings of the fluorescing groups. On the other hand, rates of dynamic quenching involving molecular motions and activation processes will vary with temperature. knr may therefore b...

show abstract

Proposed temperature-dependent conformational transition in D-amino acid oxidase: a differential scanning microcalorimetric study.

Cited by 25 publications

References 11 publications

Evaluation of the thermal coefficient of the resistance to fluorophore rotation in model membranes

Evaluation of the thermal coefficient of the resistance to fluorophore rotation in model membranes

Deflavination and reconstitution of flavoproteins

Spurious conformational transitions in proteins?

Contact Info

Product

Resources

About