Site directed modification of beta-lactoblobulin and its effect on in-vitro digestibility

Richard Owusu-Apenten

Research output: Chapter in Book/Report/Conference proceedingConference contribution

Abstract

Digestibility and amino acid composition are important considerations for assessing protein nutrient value in terms of Protein Digestibility Corrected Amino Acid Scores (PDCAAS). We hypothesize that quaternary structure adversely affects protein digestibility and other nutritional characteristics such as allergenicity and gastrointestinal toxicity. The objective of this ongoing study is to determine the effect of site directed chemical modification on beta-lactoglobulin (BLG) in vitro digestibility by pepsin. BLG was subjected to highly specific chemical modifications at two side-chain groups located at the dimer subunit interface. Cysteine 121 was modified using iodine/ potassium iodide assisted oxidation in the presence of thiols including, cysteine, mecaptoethanol, 2-mecaptoethanoic acid and cysteamine. In another series of studies, BLG was reacted with diacetyl and other arginine-specific reagents. Cys 121 or Arg 40 modified varieties (BLG121 & BLG40) were tested for stability using differential scanning calorimetry (DSC). Pepsin digestibility was measured using quantitative sodium dodecylsulfate polyacrylamide gel electrophoresis (QSDS-PAGE). The results showed that BLG121 and BLG40 varieties had pepsin digestibility values of 70-100% compared to 0% for unmodified BLG. Analysis by DSC revealed that chemical modification produced structural alterations ranging from, stabilization, destabilization, and complete loss of native structure. All modified BLG121 and BLG40 varieties showing reduced stability also showed improved in vitro digestibility. However, some stabilized BLG121 and BLG40 varieties also exhibited improved digestibility. No simple relationship could be demonstrated between protein stability and BLG in vitro digestibility. Reducing BLG stability should enhance protein dissociation and unfolding leading to greater susceptibility to pepsin. Some digestibility increases could be explained by changes in pepsin specificity as new chemical groups are introduced into BLG. Site directed chemical modification, of amino acid residues located at protein subunit interfaces, appears to be a useful technique for improving protein quality.
LanguageEnglish
Title of host publicationUnknown Host Publication
Number of pages1
Publication statusPublished - 2003
Event2003 IFT Annual Meeting, July 12 - 16, Chicago, IL - Chicago, IL
Duration: 1 Jan 2003 → …

Conference

Conference2003 IFT Annual Meeting, July 12 - 16, Chicago, IL
Period1/01/03 → …

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in vitro digestibility
beta-lactoglobulin
pepsin
digestibility
differential scanning calorimetry
digestible protein
cysteine
proteins
nutritive value
potassium iodide
cysteamine
allergenicity
diacetyl
amino acids
protein subunits
thiols
iodine
amino acid composition
arginine
polyacrylamide gel electrophoresis

Cite this

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title = "Site directed modification of beta-lactoblobulin and its effect on in-vitro digestibility",
abstract = "Digestibility and amino acid composition are important considerations for assessing protein nutrient value in terms of Protein Digestibility Corrected Amino Acid Scores (PDCAAS). We hypothesize that quaternary structure adversely affects protein digestibility and other nutritional characteristics such as allergenicity and gastrointestinal toxicity. The objective of this ongoing study is to determine the effect of site directed chemical modification on beta-lactoglobulin (BLG) in vitro digestibility by pepsin. BLG was subjected to highly specific chemical modifications at two side-chain groups located at the dimer subunit interface. Cysteine 121 was modified using iodine/ potassium iodide assisted oxidation in the presence of thiols including, cysteine, mecaptoethanol, 2-mecaptoethanoic acid and cysteamine. In another series of studies, BLG was reacted with diacetyl and other arginine-specific reagents. Cys 121 or Arg 40 modified varieties (BLG121 & BLG40) were tested for stability using differential scanning calorimetry (DSC). Pepsin digestibility was measured using quantitative sodium dodecylsulfate polyacrylamide gel electrophoresis (QSDS-PAGE). The results showed that BLG121 and BLG40 varieties had pepsin digestibility values of 70-100{\%} compared to 0{\%} for unmodified BLG. Analysis by DSC revealed that chemical modification produced structural alterations ranging from, stabilization, destabilization, and complete loss of native structure. All modified BLG121 and BLG40 varieties showing reduced stability also showed improved in vitro digestibility. However, some stabilized BLG121 and BLG40 varieties also exhibited improved digestibility. No simple relationship could be demonstrated between protein stability and BLG in vitro digestibility. Reducing BLG stability should enhance protein dissociation and unfolding leading to greater susceptibility to pepsin. Some digestibility increases could be explained by changes in pepsin specificity as new chemical groups are introduced into BLG. Site directed chemical modification, of amino acid residues located at protein subunit interfaces, appears to be a useful technique for improving protein quality.",
author = "Richard Owusu-Apenten",
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booktitle = "Unknown Host Publication",

}

Owusu-Apenten, R 2003, Site directed modification of beta-lactoblobulin and its effect on in-vitro digestibility. in Unknown Host Publication. 2003 IFT Annual Meeting, July 12 - 16, Chicago, IL, 1/01/03.

Site directed modification of beta-lactoblobulin and its effect on in-vitro digestibility. / Owusu-Apenten, Richard.

Unknown Host Publication. 2003.

Research output: Chapter in Book/Report/Conference proceedingConference contribution

TY - GEN

T1 - Site directed modification of beta-lactoblobulin and its effect on in-vitro digestibility

AU - Owusu-Apenten, Richard

PY - 2003

Y1 - 2003

N2 - Digestibility and amino acid composition are important considerations for assessing protein nutrient value in terms of Protein Digestibility Corrected Amino Acid Scores (PDCAAS). We hypothesize that quaternary structure adversely affects protein digestibility and other nutritional characteristics such as allergenicity and gastrointestinal toxicity. The objective of this ongoing study is to determine the effect of site directed chemical modification on beta-lactoglobulin (BLG) in vitro digestibility by pepsin. BLG was subjected to highly specific chemical modifications at two side-chain groups located at the dimer subunit interface. Cysteine 121 was modified using iodine/ potassium iodide assisted oxidation in the presence of thiols including, cysteine, mecaptoethanol, 2-mecaptoethanoic acid and cysteamine. In another series of studies, BLG was reacted with diacetyl and other arginine-specific reagents. Cys 121 or Arg 40 modified varieties (BLG121 & BLG40) were tested for stability using differential scanning calorimetry (DSC). Pepsin digestibility was measured using quantitative sodium dodecylsulfate polyacrylamide gel electrophoresis (QSDS-PAGE). The results showed that BLG121 and BLG40 varieties had pepsin digestibility values of 70-100% compared to 0% for unmodified BLG. Analysis by DSC revealed that chemical modification produced structural alterations ranging from, stabilization, destabilization, and complete loss of native structure. All modified BLG121 and BLG40 varieties showing reduced stability also showed improved in vitro digestibility. However, some stabilized BLG121 and BLG40 varieties also exhibited improved digestibility. No simple relationship could be demonstrated between protein stability and BLG in vitro digestibility. Reducing BLG stability should enhance protein dissociation and unfolding leading to greater susceptibility to pepsin. Some digestibility increases could be explained by changes in pepsin specificity as new chemical groups are introduced into BLG. Site directed chemical modification, of amino acid residues located at protein subunit interfaces, appears to be a useful technique for improving protein quality.

AB - Digestibility and amino acid composition are important considerations for assessing protein nutrient value in terms of Protein Digestibility Corrected Amino Acid Scores (PDCAAS). We hypothesize that quaternary structure adversely affects protein digestibility and other nutritional characteristics such as allergenicity and gastrointestinal toxicity. The objective of this ongoing study is to determine the effect of site directed chemical modification on beta-lactoglobulin (BLG) in vitro digestibility by pepsin. BLG was subjected to highly specific chemical modifications at two side-chain groups located at the dimer subunit interface. Cysteine 121 was modified using iodine/ potassium iodide assisted oxidation in the presence of thiols including, cysteine, mecaptoethanol, 2-mecaptoethanoic acid and cysteamine. In another series of studies, BLG was reacted with diacetyl and other arginine-specific reagents. Cys 121 or Arg 40 modified varieties (BLG121 & BLG40) were tested for stability using differential scanning calorimetry (DSC). Pepsin digestibility was measured using quantitative sodium dodecylsulfate polyacrylamide gel electrophoresis (QSDS-PAGE). The results showed that BLG121 and BLG40 varieties had pepsin digestibility values of 70-100% compared to 0% for unmodified BLG. Analysis by DSC revealed that chemical modification produced structural alterations ranging from, stabilization, destabilization, and complete loss of native structure. All modified BLG121 and BLG40 varieties showing reduced stability also showed improved in vitro digestibility. However, some stabilized BLG121 and BLG40 varieties also exhibited improved digestibility. No simple relationship could be demonstrated between protein stability and BLG in vitro digestibility. Reducing BLG stability should enhance protein dissociation and unfolding leading to greater susceptibility to pepsin. Some digestibility increases could be explained by changes in pepsin specificity as new chemical groups are introduced into BLG. Site directed chemical modification, of amino acid residues located at protein subunit interfaces, appears to be a useful technique for improving protein quality.

M3 - Conference contribution

BT - Unknown Host Publication

ER -