Hydrogen Exchange at Carbon-Hydrogen Sites during Acid or Alkaline Treatment of Proteins

Hill, Julie; Leach, Sydney

doi:10.1021/bi00900a003

Cited by 30 publications

(11 citation statements)

References 12 publications

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“…A small amount of proton exchange occurs at alanine and leucine during acid hydrolysis of some proteins (16). An enzymatic hydrolysis procedure was developed for collagen to avoid artifacts in the determination of the percent incorporation of 2H which may be introduced by this exchange.…”

Section: Measurement Ofpercent Incorporation Of2hmentioning

confidence: 99%

Deuterium nuclear magnetic resonance of specifically labeled native collagen. Investigation of protein molecular dynamics using the quadrupolar echo technique

Jelinski¹,

Sullivan²,

Batchelder³

et al. 1980

Biophysical Journal

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Collagen was labeled with [3,3,3-d3]alanine and with [d10]leucine via tissue culture. 2H nuclear magnetic resonance (NMR) spectra were obtained of collagen in solution and as fibrils using the quadrupolar echo technique. The 2H NMR data for [3,3,3-d3]alanine-labeled collagen fibrils were analyzed in terms of a model for motion in which the molecule is considered to jump between two sites, separated azimuthally by an angle 2 delta, in a time which is rapid compared with the residence time in both sites. The data suggest that the molecule undergoes reorientation over an angle, 2 delta, of approximately 30 degrees in the fibrils, and that the average angle between the alanine C alpha--C beta bond axis and the long axis of the helix is approximately 75 degrees. Reorientation is possibly segmental. The T2 for [3,3,3-d3]alanine-labeled collagen fibrils was estimated to be 105 mus. The 2H NMR data for the methyl groups of [d10]leucine-labeled collagen were analyzed qualitatively. These data established that for collagen in solution and as fibrils, rotation occurs about the leucine side-chain bonds, in addition to threefold methyl rotation and reorientation of the peptide backbone. The T2 for the methyl groups of leucine-labeled collagen is estimated to be approximately 130 mus. Taken together, these data provide strong evidence that both polypeptide backbone reorientation and amino acid side-chain motion occur in collagen molecules in the fibrils. Stabilizing interactions that determine fibril structure must therefore depend upon at least two sets of contacts in any given local region.

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Section: Measurement Ofpercent Incorporation Of2hmentioning

confidence: 99%

Deuterium nuclear magnetic resonance of specifically labeled native collagen. Investigation of protein molecular dynamics using the quadrupolar echo technique

Jelinski¹,

Sullivan²,

Batchelder³

et al. 1980

Biophysical Journal

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“…Recent investigations have focused on the racemization of amino acid residues, a process that may also be nutritionally detrimental (Hayase et al, 1973(Hayase et al, ,1975(Hayase et al, ,1979Masters and Friedman, 1979;Friedman et al, 1981;Friedman and Masters, 1982;Liardon and Hurrell, 1983;Liardon and Ledermann, 1984;Jenkins et al, 1984). Amino acid racemization occurs under various conditions of temperature, pH, and water activity (Hill and Leach, 1964;Ikawa, 1964;Manning, 1970;Hayase et al, 1975;Liardon and Hurrell, 1983) and might be activated by other food constituents like carbohydrates or lipids (Zumberge, 1979;Hayase et al, 1979). An understanding of the factors influencing amino acid racemization in protein under food-processing conditions may contribute to the production of foods with optimal nutritional value and safety.…”

Section: Introductionmentioning

confidence: 99%

“…The conversion of L amino acids in food proteins into D isomers generates nonutilizable forms of amino acids, creates peptide bonds resistant to proteolytic enzymes, and forms unnatural amino acids that may be nutritionally antagonistic or even toxic (Freimuth et al, 1978;Masters and Friedman, 1980;Bunjapamai et al, 1982;Tovar and Schwass, 1983;Friedman and Gumbmann, 1984). On the other hand, this reaction proceeds most readily a t basic pH (Tannenbaum et al, 1970;Provansal et al, 1975;Liardon and Hurrell, 1983), and factors influencing the racemization rate of several amino acids under these conditions have been studied (Hill andLeach, 1964, Masters andFriedman, 1979;Friedman et al, 1981;Friedman and Masters, 1982). Amino acid racemization occurs under various conditions of temperature, pH, and water activity (Hill and Leach, 1964;Ikawa, 1964;Manning, 1970;Hayase et al, 1975;Liardon and Hurrell, 1983) and might be activated by other food constituents like carbohydrates or lipids (Zumberge, 1979;Hayase et al, 1979).…”

Section: Introductionmentioning

confidence: 99%

Racemization kinetics of free and protein-bound amino acids under moderate alkaline treatment

Liardon¹,

Ledermann²

1986

J. Agric. Food Chem.

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557reagents, loss in activity on reduction by mercaptoethanol, and electrophoretic studies confirm that the new inhibitor is proteinaceous in nature.We could not unequivocally demonstrate the formation of an amylaseinhibitor complex during our studies. When inhibitor and human pancreatic amylase were chromatographed on Red-Sepharose, all the protein was eluted from the column whereas the native inhibitor was bound tightly to the immobilized ligand. While it should be expected that the inhibitor-enzyme complex would be eluted from Sephadex G-100 column earlier than the free enzyme and inhibitor, in our studies such a protein peak could not be identified. Instead, the inhibitor enzyme mixture was eluted in two protein peaks. The first peak with a V, value corresponding to the native inhibitor had measurable amylase activity. The second peak corresponding to native human pancreatic amylase had neither amylase activity nor inhibitory activity. However, treatment of this fraction with mercaptoethanol resulted in the appearance of amylase activity. These data suggest that the two protein peaks obtained during gel chromatography of amylaseinhibitor mixture represent two types of complexes. The fraction with amylase activity eluted first during gel chromatography could represent a complex formed due to loose interaction of inhibitor and enzyme dissociating under assay conditions exhibiting amylase activity. The second protein fraction may represent a complex formed by strong noncovalent interaction between the inhibitor and amylase, which exhibited amylase activity on inactivation of the inhibitor by treatment with mercaptoethanol. Further studies are needed to substantiate this view. Registry No. a-Amylase, 9000-90-2. Mundy, J.; Hejgaard, J.; Svendsen, I. FEBS Lett. 1984,167, 210. Petrucci, T.; Rab, A.; Tomasi, M.; Silano, V. Biochem. Biophys. Shainkin, R.; Birk, Y.Amino acid racemization was measured in a-lactalbumin, 0-lactoglobulin, a-casein, lysozyme, and BSA and in a free amino acid mixture exposed to pH 9 at 83 "C for times ranging from 0.5 to 24 or 96 h.Inversion rate constants were determined for 14 to 15 amino acids in the six models. Conclusions derived from these data were as follows: (1) Free amino acids racemize about 10 times more slowly than bound residues.(2) The main driving force in free amino acid racemization is the electron-withdrawing ability of the side chain (a* constant), except for Asp, the inversion of which seems to involve an intramolecular assistance effect.(3) The racemization of bound residues is governed by both amino side chain effects and protein-related factors. (4) Bound amino acid racemization is also affected by alkali-induced denaturation of the proteins (e.g., partial hydrolysis).

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“…Amino acid residues in proteins undergo racemization in alkaline solution more readily than do the free amino acids. These racemization reactions have been known for a long time (Dakin, 1912; Levene and Bass, 1928) and occur at different rates depending on the particular amino acid residue and experimental conditions (Neuberger, 1948; Hill and Leach, 1964;Pickering and Li, 1964;Pollock and Frommhagen, 1968;Tannenbaum et al, 1970).…”

mentioning

confidence: 99%

Reactions of phosphoproteins in alkaline solutions

Sen¹,

Gonzalez-Flores²,

Feeney³

et al. 1977

J. Agric. Food Chem.

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Phosvitin, an egg yolk protein of mol wt 35 500 with 10% phosphate in the form of O-phosphoserine residues (reported value of 119 phosphoserine and one phosphothreonine residues), was used in model systems to investigate the effects of various experimental conditions on the rates of ß elimination of phosphate from the phosphoserine residues and addition of indogenous nucleophiles to the double bond of the dehydroalanines formed. At low concentrations of phosvitin (1-10 X 6 M) the rates were independent of phosvitin concentration but directly dependent on the hydroxide ion concentration of the solution. The activation energies of the ß-elimination and addition reactions were 20.2 and 25.0 kcal/mol, respectively. The rate of the /3-elimination reaction was increased at increased ionic strengths and was markedly increased in the presence of CaCl2. At 1.12 X 10"3 M CaCl2, the rate of ß elimination was 20 times faster than in the absence of CaCl2. The rate of the addition reaction was not significantly affected by CaCl2 or ionic strength.

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Hydrogen Exchange at Carbon-Hydrogen Sites during Acid or Alkaline Treatment of Proteins

Cited by 30 publications

References 12 publications

Deuterium nuclear magnetic resonance of specifically labeled native collagen. Investigation of protein molecular dynamics using the quadrupolar echo technique

Deuterium nuclear magnetic resonance of specifically labeled native collagen. Investigation of protein molecular dynamics using the quadrupolar echo technique

Racemization kinetics of free and protein-bound amino acids under moderate alkaline treatment

Reactions of phosphoproteins in alkaline solutions

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