Ka. Johnston et al., PROTEASE ACTIVITIES IN THE LARVAL MIDGUT OF HELIOTHIS-VIRESCENS - EVIDENCE FOR TRYPSIN AND CHYMOTRYPSIN-LIKE ENZYMES, Insect biochemistry and molecular biology, 25(3), 1995, pp. 375-383
Protease activities in the midgut contents of larvae of the tobacco bu
dworm, Heliothis virescens, were investigated, Two major activities we
re present: (i) a trypsin-like enzyme, hydrolysing the synthetic subst
rate N-benzoyl-arginine p-nitroanilide (BApNA), strongly inhibited by
N-p-tosyl-lysine chloroketone (TLCK) and not inhibited by chymostatin
at concentrations where this inhibitor is chymotrypsin-specific; (ii)
an enzyme which hydrolysed synthetic chymotrypsin substrates containin
g more than one amino acid, but not N-benzoyl-tyrosine p-nitroanilide
and was strongly inhibited by chymostatin, The latter activity was con
sidered to represent a hitherto ill-characterised family of insect chy
motrypsins, which differ: from their mammalian counterparts in activit
y towards synthetic substrates, Both activities had strongly alkaline
pH optima, in the pH range 10-11, but were shown to be due to distinct
proteases, Low levels of elastase-like activity (hydrolysing succinyl
(alanine), p-nitroanilide) were also detected, The two major enzyme a
ctivities were shown to account for almost all the protease activity o
f gut contents towards protein substrates, The proteases showed differ
ent sensitivities to inhibition by plant protein protease inhibitors,
which were effective in protecting exogenous proteins from digestion b
y gut extracts.