The Use of an Escherichia coli Lys- Auxotroph to Assay Nutritionally Available Lysine in Biological Materials

A new bacterial method for determining amino acids in protein foods is described. Instead of the 'natural'microbial auxotrophs e.g. Tetrahymena, Streptococcus , and Leuconostoc , currently used for such assays, an 'artificial'mutant is used, viz. an auxotroph of Escherichia coli . Test proteins (Bovine serum albumin, legume and maize meals) were predigested with a mixture of pronase and intestinal peptidases, the efficiency and extent of proteolysis being monitored by pH stat titration. Final digests were examined by Sephadex gel filtration to ensure that all protein cleavage products were small enough to pass through the E. coli cell wall and to reach its cyto-plasmic amino acid and peptide permeases. The lysine content of the meals, as determined from the growth of an E. coli lysine auxotroph upon the digests, was found to be greater than 90° of the lysine determined chemically in acid hydrolysates. Practical and theoretical advantages of using this latter type of bacterium rather than the fastidious species are discussed. In addition, the particular value of using an intestinal bacterium like E. coli to assay nutritional availability of amino acids is considered in relation to its normal utilization of digested protein foods in vivo , and the similarities between its amino acid and peptide permeases and those of the intestine.