The use ofEscherichia coli mutants to measure the bioavailability of essential amino acids in foods

Abstract

The beta-galactosidase-based assay for lysine developed by Tuffnell & Payne was used to measure the bioavailabilities of cyst(e)ine, methionine, threonine and tryptophan in pronase digests of 17 foods. The digests were assayed by estimating the beta-galactosidase synthesis responses of five Escherichia coli mutants, each requiring one of the respective amino acids for protein synthesis. Deletion mutants were used whenever possible in order to ensure strain stability. Single digests of each food were assayed with 3 or 4 separate cultures of each mutant and the results were compared with those from the corresponding chemical assay. Omitting the anomalously low values for one food, the rank correlation coefficients of the bio- and chemo-assay values were 0.61 (cysteine), 0.91 (lysine), 0.95 (methionine), 0.64 (threonine) and 0.85 (tryptophan). Mean (+/- S.D.) relative amino acid bioavailabilities (casein = 100%) for the 17 foods were: cysteine, 53 +/- 23; lysine, 90 +/- 10; methionine, 95 +/- 18; threonine, 89 +/- 13; and tryptophan, 89 +/- 25. The cysteine mutant appeared to give unusually low bioavailability values except for the milk products. These amino acid mutants afford a rapid method for assaying the bioavailabilities of at least four of the five amino acids studied.

Optimisation of enzyme treatment for the degradation of feed proteins for an Escherichia coli auxotroph lysine availability assay

“…Casein and whole dried egg are both highly available protein sources, with casein considered to be completely available. 16,30 Poultry by-product meal, however, has lower and more highly variable amino acid digestibility than casein or egg. 27,32 When comparing enzyme treatment only on the protein sources, protease as a single-enzyme digest was as complete at digesting the protein sources as a multiple-enzyme treatment.…”

“…Pepsin, chymotrypsin, trypsin, peptidase and protease all have been utilised. 11,13,16 Based upon a mixture of protease and peptidase producing the most extensive digest, this enzyme combination has been recommended for use. 11 However, the duration of enzyme treatment of the protein sources for E coli has varied in studies from 2 to 16 h. 11,16 The duration of the enzyme hydrolysis can affect the total quantity of amino acids and peptides released.…”

“…11,13,16 Based upon a mixture of protease and peptidase producing the most extensive digest, this enzyme combination has been recommended for use. 11 However, the duration of enzyme treatment of the protein sources for E coli has varied in studies from 2 to 16 h. 11,16 The duration of the enzyme hydrolysis can affect the total quantity of amino acids and peptides released. 11,17,18 Therefore varying the length of enzyme incubation with the protein source and using different mixtures of enzymes for protein digestion may yield better correlation with animal data, thus replacing the need for complete reliance on animal trials 19 and allowing for standardisation.…”

Potential for Development of an Escherichia coli—Based Biosensor for Assessing Bioavailable Methionine: A Review

“…Hitchens et al [45] demonstrated that E. coli GUC41 could grow on DL-methionine sulfoxide but not on DL-methionine sulfone. The microbiological assay was as accurate as the chemical assay with high correlation coefficients between the two.…”

Generation of a green fluorescent protein gene chromosomal insertion containing Escherichia coli strain for gene induction-based quantification of bioavailable lysine

“…Several attempts have been made to develop rapid methods to estimate lysine bioavailability in foods and feeds, including chemical methods [2,3] and microbiological assays [4,5]. Even though the analytical chemical methods are more rapid when compared to the conventional animal bioassays, they fail to measure lysine availability accurately [6,7].…”