Jr. Reid et al., SPECIFICITY OF HYDROLYSIS OF BOVINE KAPPA-CASEIN BY CELL ENVELOPE-ASSOCIATED PROTEINASES FROM LACTOCOCCUS-LACTIS STRAINS, Applied and environmental microbiology, 60(3), 1994, pp. 801-806
The cell envelope-associated proteinases from Lactococcus lactis subsp
. cremoris H2 (a P-I-type proteinase-producing strain) and SK11 (a P-I
II-type proteinase-producing strain) both actively hydrolyze the kappa
-casein component of bovine milk but with significant differences in t
he specificity of peptide bond hydrolysis. The peptide bonds Ala-23-Ly
s-24, Leu-32-Ser-33, Ala-71-Gln-72, Leu-79-Ser-80, Met-95-Ala-96, and
Met-106-Ala-107 were cleaved by both proteinase types, although the re
lative rates of hydrolysis at some of these sites were quite different
for the two proteinases. Small histidine-rich peptides were formed as
early products of the action of the cell envelope-associated proteina
ses on kappa-casein, implicating this casein as a possible significant
source of histidine, which is essential for starter growth. The major
difference between the two proteinase types in their action on kappa-
casein was in their ability to cleave bonds near the C-terminal end of
the molecule. The bond Asn-160-Thr-161 and, to a lesser extent, the b
ond Glu-151-Val-152 were very rapidly cleaved by the P-III-type protei
nase, whereas hydrolysis of these bonds by the P-I-type proteinase was
barely detectable (even after 24 h of digestion). Differential hydrol
ysis of kappa-casein at these sites by the two different proteinase ty
pes resulted in the formation of distinctive, high-M(r) products detec
table by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Th
e kappa-casein cleavage patterns revealed by sodium dodecyl sulfate-po
lyacrylamide gel electrophoresis analysis provide a diagnostic basis f
or recognizing differences in proteinase specificity and have been use
d to identify P-I- or P-III-type and P-I/P-III intermediate-type prote
inases from several different lactococcal strains.