Isolement et determination de la composition qualitative de peptides issus de la caséine, stimulant la croissance de Streptococcus thermophilus
Authors: Desmazeaud Michel J. , Hermier Jean H. 
Published: March 2005

In order to ascertain the physiological role of the peptides which enhanced the growth of Streptococcus thermophilus in milk, fourteen stimulating peptides were isolated and purified and their qualitative amino‐acid composition was determined. The purification process was the following: after hydrolysis at pH 7.5 of whole casein by the neutral protease from Micrococcus caseolyticus the stimulating peptides were fractionated by gel chromatography (Bio‐Gel P6), by column chromatography on cation‐exchange resins AG SOW X2 (0.05 M pyridine‐formate buffer at pH 3.10 to 1.5M pyridine‐acetate buffer at pH 5.10). Further purification was made by column chromatography on anion‐exchange resins AG1 X2 (0.1 M pyridine‐acetate buffer of pH 7.5 to 6 M acetic acid with a concave gradient system) and by paper chromatography (solvent: butanol–acetic acid–water, 4:1:5, v/v/v) and high tension electrophoresis (30 min; 60 V/cm).

Gel filtration of enzymic digest of the whole casein led to six peptide fractions. Three fractions were stimulatory and exhibited a mean molecular weight of (B) 2500, (C) 1500, (D) 1000. Fraction C was the most stimulatory. Three other fractions were inhibitory and contained the highest molecular‐weight peptides (about 5000) and peptides with high contents of aromatic residues. Ten and twenty peptidic fractions were isolated respectively from Fractions B and D by chromatography on ion‐exchange resins; but the peptides obtained after further purification were no more stimulatory. Eleven stimulatory and two inhibitory fractions were separated from Fraction C by column chromatography on cation‐exchange resins. Purified stimulatory peptides were obtained from two fractions (C1 and C42) only, after anion‐exchange resins (three peptides from Fraction C42, eleven peptides from Fraction C1).

These fourteen purified stimulatory peptides display a very variable qualitative amino‐acid composition. The polar part (Lys, Glx, Asx) was as important as the hydrophobic part (Pro, Leu, Val, Ala, Gly). The number of distinct ammo‐acid residues from one peptide was also quite variable: fourteen to four residues. Only lysyl residue was present in all peptides and tryptophan was not present. Three peptides that contained cysteine were issued from ϰ‐casein [18]; two peptides (Cla2α and C1C3) could be located respectively to Ala26… Gln52 and Ile81…Arg119 regions from bovine αs1B casein.

A complete mixture of amino acids exerted growth stimulation. The set of the ammo acids present in a stimulatory peptide was always stimulatory.

These data substantiate the following conclusions: stimulatory peptides acted as amino‐acid source; the amino acids have to be supplied in specific proportions in order not to disturb the cellular amino acids pool and the regulation of amino acid synthesis by Str. thermophilus; (b) the requirements in the peptide‐length residues and lysyl residue could be explained by a control system of peptide intake, for example, by a peptide‐transport system that would differ from that described in Escherichia coli.