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. 2000 Dec;66(12):5134-40.
doi: 10.1128/AEM.66.12.5134-5140.2000.

The autoproteolysis of Lactococcus lactis lactocepin III affects its specificity towards beta-casein

B Flambard  1 , V Juillard
Affiliations

The autoproteolysis of Lactococcus lactis lactocepin III affects its specificity towards beta-casein

B Flambard et al. Appl Environ Microbiol. 2000 Dec.

Abstract

The effect of autoproteolysis of Lactococcus lactis lactocepin III on its specificity towards beta-casein was investigated. beta-Casein degradation was performed by using either an autolysin-defective derivative of L. lactis MG1363 carrying the proteinase genes of L. lactis SK11, which was unable to transport oligopeptides, or autoproteolyzed enzyme purified from L. lactis SK11. Comparison of the peptide pools by high-performance liquid chromatography analysis revealed significant differences. To analyze these differences in more detail, the peptides released by the cell-anchored proteinase were identified by on-line coupling of liquid chromatography to mass spectrometry. More than 100 oligopeptides were released from beta-casein by the cell-anchored proteinase. Analysis of the cleavage sites indicated that the specificity of peptide bond cleavage by the cell-anchored proteinase differed significantly from that of the autoproteolyzed enzyme.

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Figures

FIG. 1
FIG. 1
TFA (1%)-soluble peptides released from β-casein by autoproteolyzed lactocepin III (bottom line) and cell-anchored lactocepin III (top line). Autoproteolyzed lactocepin III was obtained by incubating L. lactis SK11 cells in a Ca2+-free buffer and was further purified by ion-exchange chromatography.
FIG. 2
FIG. 2
TFA (1%)-soluble peptides released from β-casein by autoproteolyzed lactocepin III in the presence of proteinase-negative strain L. lactis GF100. Autoproteolyzed lactocepin III was obtained by incubating L. lactis SK11 cells in a Ca2+-free buffer and was further purified by ion-exchange chromatography.
FIG. 3
FIG. 3
Localization of the peptides released by the action of cell-anchored lactocepin III on β-casein (A2 variant). The dotted arrows indicate peptides identified as peptides that were also released by autoproteolyzed lactocepin III (data from reference 38).
See this image and copyright information in PMC

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