A new
protease has been purified to homogeneity from rat submandibular gland homogenate by using
DEAE-Sephadex chromatography, chromatofocusing,
aprotinin-
Sepharose affinity chromatography, and high-performance liquid chromatography. The
enzyme has been named
esterase B, since it represents the second major esterolytic peak on
DEAE-Sephadex chromatography of submandibular gland homogenate. It is an acidic
protein (pI = 4.45) with an apparent molecular weight of 27 000. It is heat-stable and has an optimum pH of 9.5.
Esterase B hydrolyzed the synthetic substrates tosyl-
L-arginine methyl ester and
Val-Leu-Arg-p-nitroanilide (
S2266). It also cleaved dog plasma
kininogen to produce a
kinin, identified as
bradykinin on reverse-phase high-performance liquid chromatography.
Esterase B, however, is only a weak
kininogenase, since it had only 5% of the
kininogenase activity of equimolar concentrations of
glandular kallikrein and had no effect on rat mean blood pressure or on the isolated rat uterus.
Esterase B activated
plasminogen and had caseinolytic activity. It was inhibited by
aprotinin, soybean
trypsin inhibitor, lima bean
trypsin inhibitor,
phenylmethanesulfonyl fluoride,
antipain,
leupeptin, and p-tosyl-
L-lysine chloromethyl
ketone. On double immunodiffusion, when reacted with
kallikrein and
tonin antisera,
esterase B showed partial identity with
kallikrein but not with
tonin. On immunoelectrophoresis against
kallikrein antisera,
esterase B formed a precipitin
arc at a position different from that of
kallikrein.
Esterase B appears to be a
trypsin-like serine protease having some homology with
glandular kallikrein.