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Sequence analysis and molecular characterization of Clonorchis sinensis hexokinase, an unusual trimeric 50-kDa glucose-6-phosphate-sensitive allosteric enzyme.
[cholangiocarcinoma]
Clonorchiasis
,
which
is
induced
by
the
infection
of
Clonorchis
sinensis
(
C
.
sinensis
)
,
is
highly
associated
with
cholangiocarcinoma
.
Because
the
available
examination
,
treatment
and
interrupting
transmission
provide
limited
opportunities
to
prevent
infection
,
it
is
urgent
to
develop
integrated
strategies
to
prevent
and
control
clonorchiasis
.
Glycolytic
enzymes
are
crucial
molecules
for
trematode
survival
and
have
been
targeted
for
drug
development
.
Hexokinase
of
C
.
sinensis
(
CsHK
)
,
the
first
key
regulatory
enzyme
of
the
glycolytic
pathway
,
was
characterized
in
this
study
.
The
calculated
molecular
mass
(
Mr
)
of
CsHK
was
50
.
0
kDa
.
The
obtained
recombinant
CsHK
(
rCsHK
)
was
a
homotrimer
with
an
Mr
of
approximately
164
kDa
,
as
determined
using
native
PAGE
and
gel
filtration
.
The
highest
activity
was
obtained
with
50
mM
glycine-
NaOH
at
pH
10
and
100
mM
Tris-
HCl
at
pH
8
.
5
and
10
.
The
kinetics
of
rCsHK
has
a
moderate
thermal
stability
.
Compared
to
that
of
the
corresponding
negative
control
,
the
enzymatic
activity
was
significantly
inhibited
by
praziquantel
(
PZQ
)
and
anti-r
CsHK
serum
.
rCsHK
was
homotropically
and
allosterically
activated
by
its
substrates
,
including
glucose
,
mannose
,
fructose
,
and
ATP
.
ADP
exhibited
mixed
allosteric
effect
on
rCsHK
with
respect
to
ATP
,
while
inorganic
pyrophosphate
(
PPi
)
displayed
net
allosteric
activation
with
various
allosteric
systems
.
Fructose
behaved
as
a
dose-dependent
V
activator
with
the
substrate
glucose
.
Glucose-
6
-
phosphate
(
G
6
P
)
displayed
net
allosteric
inhibition
on
rCsHK
with
respect
to
ATP
or
glucose
with
various
allosteric
systems
in
a
dose-independent
manner
.
There
were
differences
in
both
mRNA
and
protein
levels
of
CsHK
among
the
life
stages
of
adult
worm
,
metacercaria
,
excysted
metacercaria
and
egg
of
C
.
sinensis
,
suggesting
different
energy
requirements
during
different
development
stages
.
Our
study
furthers
the
understanding
of
the
biological
functions
of
CsHK
and
supports
the
need
to
screen
for
small
molecule
inhibitors
of
CsHK
to
interfere
with
glycolysis
in
C
.
sinensis
.
Diseases
Validation
Diseases presenting
"mixed allosteric effect on rcshk"
symptom
cholangiocarcinoma
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