Fast protein liquid chromatography purification of hydrophobic fraction of bovine milk proteose-peptone and characterization by bidimensional electrophoresis

Girardet, Jean‐Michel; Mati, Abderrahmane; Sanogo, T.; Etienne, L.; Linden, G.

doi:10.1017/s0022029900033537

Cited by 20 publications

(19 citation statements)

References 33 publications

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“…NHF, LHF and HHF were identical to fractions l, 1I+1I1+IV and V respectively, isolated in our earlier work (Girardet et al, 1991). So, NHF was composed by hydrophilic peptides or polypeptides.…”

Section: Distribution Of Activity Among Proteosepeptone Fractionssupporting

confidence: 72%

“…Proteose-peptone fractionation was pertermed using a TSK-Phenyl-5PW column connected to the FPLC system (Girardet et al, 1991). Sample (150-200 mg) was loaded on the column equilibrated in 0.8 mol/l-NaH2P04 buffer (pH 6.8) at 25°C.…”

Section: Fraetionation Of Proteose-peptonementioning

confidence: 99%

“…The first includes polypeptides resulting from proteolysis of casein by endogenous proteinase in milk among which p-CN-5P (f1-1 05/1 07; 12 441 Da), p-CN-4P (f1-28; 3478 Da), and p-CN-1 P (f29-1 05/1 07; 8981 Da) are N-terminal fragments of p-casein (Eigel et al, 1984). The second group can be separated from the first by hydrophobic interaction liquid chromatography (Pâquet et al, 1988;Girardet et al, 1991) and contains three principal glycoproteins with apparent M r of 11 000, 19 000, and 29 000 Da. The primary structure of component 3 corresponding to the 29 OOO-Da glycoprotein has been recently determined (Serensen and Petersen, 1993) and shown to contain 135 amine acid residues.…”

Section: Introductionmentioning

confidence: 99%

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Growth promotion of Bifidobacterium animalis by bovine milk proteose-peptone

Etienne¹,

Girardet²,

Linden³

1994

Lait

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Summary -The industrial strain Bifidobacterium animalis was used as assay organism to evaluate bifidobacterial growth-promoting activity of bovine milk proteose-peptone. This proved to be a better growth-promoting factor than bovine casein. The bifidogenic activity was found mainly in the proteose-peptone hydrophobie fraction containing component 3, although the glycan moiety was a weak growth-promoter. Proteose-peptone digests by various proteolytic enzymes caused great enhancement of B animalis growth, particularly the Pronase digest. Size-exclusion chromatography of digests showed that the more active peptides had a molecular mass distribution of 1000 to 5000 Da.

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Section: Distribution Of Activity Among Proteosepeptone Fractionssupporting

confidence: 72%

Section: Fraetionation Of Proteose-peptonementioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Growth promotion of Bifidobacterium animalis by bovine milk proteose-peptone

Etienne¹,

Girardet²,

Linden³

1994

Lait

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“…-CN f(1-105/7) was purified by hydrophobic interaction FPLC on TSK-Phenyl 5PW (Pharmacial, following the methods of Paquet et al, (1988) and Girardet et al (1991).…”

Section: Biochemical Reagentsmentioning

confidence: 99%

Proteolysis of bovine β-casein by a plasmin-Sepharose conjugate

Bénali¹,

Guilbaud²,

Montagne

et al. 1994

Lait

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Summary -A plasmin-Sepharose conjugate (5.7 mg of active plasmin/g gel) was prepared by covalent immobilization of bovine plasmin (EC 3.4.21.7) on activated CH-Sepharose 48, with formation of amide bonds between primary amino groups of plasmin and carboxyl-activated functions anchored to the gel. A strong decrease (60%) of the proteolytic activity of plasmin for a chromogenic substrate was observed during dialysis performed before immobilization. However, a high binding yield and a goOO preservation of the proteolytic activity of immobilized enzyme (95%) were then observed. The reactivity of free and immobilized plasmin with bovine p-easein (P-CN) were compared. Kinetic study of p-CN proteolysis indicated a perceptible loss of enzyme activity in plasmin-Sepharose conjugate compared to free plasmin (Vmax = 3.8 10-2 and 53.5 10-2 00 unit/min, Km = 1.2 1Q-4 et 6.2 10-4 molli, respectively). Electrophoretic study of the reaction mixtures showed that the proteolysis of p-CN f(1-10517) by immobilized or free plasmin, producing p-CN f(29-1 0517), succeeds the reaction of the whole p-CN.Microparticle-enhanced nephelometric immunoassay of residual p-CN, performed in reaction supernatants, indicated there is no significant influence of p-CN fragments (P-CN f(1-1 0517) and mixture of C-terminal peptides p-CN (29-209), (106-209), (108-209)) on the proteolysis of the whole p-CN. As already observed for other proteases, an alteration of the enzyme charge during the immobilization process might be the cause of the slight modification of the catalytic activity observed in plasmin-conjugate.

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“…Le PP3 se retrouve en majorité dans ce dernier groupe de constituants (PM = 28 000 Da), alors que les composants~-CN-5P, -CN-4P et~-CN-1P sont représentés dans le second groupe (PM = 18 000 Da) par des spots à PI acides (de 4 à 5,5). L'ensemble de ces résultats concorde avec ceux obtenus par Girardet et al (1991) sur des fractions analogues de lait bovin séparées par FPLC.…”

Section: Analyses Chimiquesunclassified

Isolement et caractérisation de la fraction hydrophobe des protéose-peptones des laits bovin, ovin et caprin

Mati

Girardet

Xenakis

et al. 1991

Lait

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(Reçu le 9 octobre 1990; accepté le 15 décembre 1990) Résumé -Le protocole d'isolement des protéose-peptones, largement utilisé jusque là pour le lait de vache, a été adapté et appliqué aux laits de brebis et de chèvre. Les fractions de ces protéines contenant le composant-s sont isolées par chromatographie d'interactions hydrophobes. Les rendements d'extraction sont améliorés par optimisation .de ce protocole. Le comportement physicochimique des fractions protéiques (protéose-peptones totales et leurs fractions hydrophobes) est étudié, L'électrophorèse en gel de polyacrylamide en milieu non dissociant (PAGE native) montre une plus faible mobilité du composant-3 des laits ovin et caprin par rapport à leur homologue du lait bovin. En milieu dissociant, en présence de dodécylsulfate de sodium et du 2-mercaptoéthanol (PAGE-SDS), les profils électrophorétiques des fractions hydrophobes ont en commun 2 bandes principales correspondant à des glycoprotéines de 18 000 et 28 000 Da. Ces polypeptides sont impliqués dans les agrégats de hauts poids moléculaires mis en évidence par chromatographie de perméation sur gel (colonnes de Séphacryl S200 et Superose 12). L'électrophorèse bidimensionnelle a montré une bonne similitude entre les fractions protéiques des laits des 3 espèces, concernant leurs points isoélectriques et leurs poids moléculaires. La composition chimique (acides aminés, glucides, phosphore) a montré également des analogies entre les fractions protéose-peptones des laits provenant des espèces laitières étudiées.

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Fast protein liquid chromatography purification of hydrophobic fraction of bovine milk proteose-peptone and characterization by bidimensional electrophoresis

Cited by 20 publications

References 33 publications

Growth promotion of Bifidobacterium animalis by bovine milk proteose-peptone

Growth promotion of Bifidobacterium animalis by bovine milk proteose-peptone

Proteolysis of bovine β-casein by a plasmin-Sepharose conjugate

Isolement et caractérisation de la fraction hydrophobe des protéose-peptones des laits bovin, ovin et caprin

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