Increase in the stability of avidin produced by binding of biotin. Differential scanning calorimetric study of denaturation by heat

Donovan, John W.; Ross, Kenneth D.

doi:10.1021/bi00727a024

Cited by 127 publications

(47 citation statements)

References 27 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The obvious source of explanation of this phenomenon is the ligand. Biotin binding to apoavidin protects against the action of denaturing agents, and the T,, is increased from 85 "C to 132 "C when biotin is bound (Donovan & Ross, 1973;Green, 1975). Clearly, the binding of the natural ligand induces considerable additional stability in the protein.…”

Section: Discussionmentioning

confidence: 99%

Limited proteolysis of native proteins: The interaction between avidin and proteinase K

et al. 1995

View full text Add to dashboard Cite

Avidin is a tetramer of 16-kDa subunits that have a high affinity for biotin. Proteolysis of native apoavidin by proteinase K results in a limited attack at the loop between P-strands 3 and 4, involving amino acids 38-43. Specifically, sites of proteolysis are at Thr 40-Ser 41 and Asn 42-Glu 43. The limited proteolysis results in an avidin product that remains otherwise intact and which has enhanced binding for 4'-hydroxyazobenzene-2-benzoic acid (HABA), a chromogenic reporter that can occupy the biotin-binding site. Saturation of the biotin-binding site with the natural ligand protects avidin from proteolysis, but saturation with HABA enhances the rate of proteolysis of the same site.Analysis of the three-dimensional structures of apoavidin and holoavidin reveals that the 3-4 loop is accessible to solvent and scores highly in an algorithm developed to identify sites of proteolytic attack. The structure of holoavidin is almost identical to the apoprotein. In particular, the 3-4 loop has the same structure in the apo and holo forms, yet there are marked differences in proteolytic susceptibility of this region. Evidence suggests that the 3-4 loop is rather mobile and flexible in the apoprotein, and that it becomes constrained upon ligand binding. In one crystal structure of the apoprotein, this loop appears constrained by contacts with symmetry-related molecules. Structural analyses suggest that the "lid" to the biotin-binding site, formed by the 3-4 loop, is displaced and made more accessible by HABA binding, thereby enhancing its proteolytic susceptibility.

show abstract

Section: Discussionmentioning

confidence: 99%

Limited proteolysis of native proteins: The interaction between avidin and proteinase K

et al. 1995

View full text Add to dashboard Cite

show abstract

“…The rate constant for denaturation was calculated at various temperatures and heating rates. 9 The vertical deflection from the baseline, at any temperature, is proportional to the rate of heat flow into the sample, dH/dt, and thus is a measure of the rate of denaturation. The relative amount of native protein present at any temperature T can be determined by measuring the area under the peak above that temperature.…”

Section: Methodsmentioning

confidence: 99%

A differential scanning calorimetric study of the stability of egg white to heat denaturation

Donovan¹,

Mapes²,

Davis³

et al. 1975

J Sci Food Agric

Self Cite

214

View full text Add to dashboard Cite

The heat denaturation of egg white and its component proteins was studied by differential scanning calorimetry. At a heating rate of 10 "C/min, egg white at pH 7 shows two major endotherms, at 65 "C and 84 "C, produced by the denaturation of conalbumin and ovalbumin, respectively. The conalbumin endotherm is increased to 70 "C by raising the pH to 9.0, or 77 "C by addition of aluminium at neutral pH. Addition of sucrose stabilises all the proteins; at 10% sucrose, all endotherms are shifted 2 "C to higher temperatures. Within experimental error, the enthalpy of denaturation of egg white equals the sum of the enthalpies of denaturation of its component proteins, and is independent of pH over the pH range 7-9.

show abstract

“…In fairly good agreement with DSC results Hegg et al, 1978) a significant rise in transverse water proton relaxation rates, proportional to the protein content of the sample, was observed in apoovotransferrin aqueous solutions (pH 7.0) after they were heated for 30 min at temperatures between 50 and 60 "C ( Figure 4). Thermal transitions observed by DSC during heating of globular protein solutions are related to protein unfolding denaturation (Donovan and Ross, 1973;Privalov and Khechinashvili, 1974;Brandts, 1969). The variation in transverse relaxation rates observed during heating conalbumin solution can therefore be associated with changes in the conformation of the protein.…”

Section: Protein C O N C E N T R a T I O N (W/w)mentioning

confidence: 99%

Low-field nuclear magnetic resonance relaxation study of the thermal denaturation of transferrins

Lambelet¹,

Ducret²,

Leuba³

et al. 1991

J. Agric. Food Chem.

View full text Add to dashboard Cite

The NMR method previously described by Lambelet et al. (J. Dairy Res. 1989,56,211-222) has been evaluated for the investigation of thermal and acid denaturation of transferrins in aqueous solution.Heating an apoovotransferrin or an apolactoferrin solution resulted in a slight increase in T1-l and an important increase in T2-1 relaxation rates within the denaturation range. The same treatment applied to corresponding iron-saturated proteins resulted in a decrease in these parameters due to modifications of the metal-protein complex accompanying the denaturation. It is proposed that these changes in NMR relaxation parameters can be used to monitor thermal denaturation of transferrins. Acid treatment led to a decrease in relaxation parameters for both apoovotransferrin and iron-saturated (down to pH 4) ovotransferrin. These variations in NMR parameters were related to protein denaturation and indicated that the protein modifications due to acid treatment were different from those occurring during heating.

show abstract

Increase in the stability of avidin produced by binding of biotin. Differential scanning calorimetric study of denaturation by heat

Cited by 127 publications

References 27 publications

Limited proteolysis of native proteins: The interaction between avidin and proteinase K

Limited proteolysis of native proteins: The interaction between avidin and proteinase K

A differential scanning calorimetric study of the stability of egg white to heat denaturation

Low-field nuclear magnetic resonance relaxation study of the thermal denaturation of transferrins

Contact Info

Product

Resources

About