1)
The
figure
below
shows
two
amino
acid
residues
in
the
active
site
of
an
enzyme.
All
non-‐hydrogen
atoms
are
displayed;
oxygen
and
nitrogen
atoms
are
labelled.
a)
Name
the
type
of
amino
acid
present
in
the
active
site.
[1
mark]
b)
What
is
meant
by
the
pKa
of
an
ionisable
group?
[4
marks]
c)
What
will
be
the
effect
of
the
close
proximity
of
these
two
side-‐chains
on
their
pKa
values?
Briefly
explain
your
answer.
[3
marks]
d)
These
active
site
residues
are
relatively
exposed
to
the
solvent,
but
if
they
were
in
a
more
hydrophobic
micro-‐environment
in
the
active
site
of
an
enzyme,
what
effect
would
this
have
on
their
pKa
values?
Briefly
explain
your
answer.
[4
marks]
e)
Cyrus
Levinthal
proposed
a
thought
experiment
to
examine
the
possible
mechanisms
by
which
proteins
fold.
Briefly
outline
Levinthal’s
argument.
Why
is
the
apparently
paradoxical
conclusion
of
this
experiment
not
observed
in
nature?
[8
marks]
2)
a)
Protein
structures
can
be
experimentally
determined
to
high
resolution
using
the
techniques
of
nuclear
magnetic
resonance
(NMR),
X-‐ray
crystallography
and
single-‐particle
averaging
using
transmission
electron
microscopy
(TEM).
Briefly
outline
the
chief
advantages
and
disadvantages
of
these
methods.
[6
marks]
b)
Why
do
the
values
for
the
phi
(ϕ)
and
psi
(ψ)
angles
of
the
polypeptide
chain
cluster
in
the
‘most
favoured’
regions
of
the
Ramachandran
plot?
How
is
this
useful
when
experimentally
determining
a
protein
structure?
[3
marks]
c)
Proteins
have
evolved
to
be
only
marginally
stable
in
their
folded
forms.
Why
is
this
an
advantage
in
the
cell?
[2
marks]
d)
Describe
how
the
redox
chemistry
of
cysteine
influences
protein
folding
in
extracellular
proteins.
[3
marks]
e)
What
is
the
amino
acid
sequence
of
this
peptide,
written
using
one-‐letter
abbreviations
in
the
conventional
orientation?
[4
marks]
f)
What
is
the
net
charge
on
the
following
peptide
in
the
cell,
assuming
a
buffered
intracellular
pH
of
7.3?
M-‐S-‐Q-‐R-‐E-‐G-‐N-‐P-‐D-‐F-‐Y-‐W-‐C-‐T-‐I-‐K-‐L-‐A-‐V
[2
marks]
3)
When
answering
the
following
questions,
remember
that
the
cellular
interior
is
buffered
at
approximately
pH
7.3,
often
referred
to
as
‘physiological
pH’.
Consider
the
following
peptide
sequence:
M-‐S-‐R-‐T-‐K-‐I-‐N-‐E-‐D-‐P-‐L-‐W-‐E-‐Y-‐S-‐G
a)
Assuming
no
influence
from
protein
tertiary
structure,
will
this
peptide
have
an
overall
charge
in
the
cell,
and
if
so,
will
it
be
positively
or
negatively
charged?
Explain
the
reasoning
behind
your
answer.
[4
marks]
b)
Now
assume
that
the
arginine
residue
is
placed
in
close
physical
proximity
to
an
aspartate
residue
due
the
context
of
the
peptide
sequence
in
a
folded
protein.
Briefly
explain
what
will
happen
to
the
pKa
value
of
the
side-‐chain
of
the
aspartate
residue,
compared
to
its
resting
value
in
solution.
How
will
this
be
different
if
the
residues
are
buried
in
a
hydrophobic
environment
within
the
folded
protein?
[4
marks]
c)
Why
is
the
maintenance
of
‘physiological’
pH
important
for
the
preservation
of
correctly
folded
proteins
within
the
cell?
[4
marks]
d)
Describe
briefly
how
the
distinctive
redox
chemistry
of
cysteine
may
influence
protein
folding
and
function
in
extracellular
proteins.
How
is
this
different
in
intracellular
proteins?
[4
marks]
e)
Describe
the
distinctive
hydrogen
bonding
patterns
found
in
the
secondary
structure
elements
of
proteins.
[4
marks]
4)a)
Draw
a
diagram
of
a
polypeptide
chain,
labelling
the
atoms
and
three
torsion
(dihedral)
angles
that
are
repeated
along
the
backbone.
[2
marks]
How
is
protein
structure
constrained
by
the
varying
degrees
of
freedom
of
rotation
around
these
three
bonds?
You
should
illustrate
your
answer
with
diagrams
as
appropriate.
[6
marks]
b)
Describe
Cyrus
Levinthal’s
thought
experiment
concerning
a
possible
mechanism
of
protein
folding.
What
does
this
experiment
tell
us?
[4
marks]
c)
Proteins
have
evolved
to
be
only
marginally
stable
in
their
folded
form.
Why?
[2
marks]
d)
Describe
the
forces
that
drive
the
folding
of
globular
proteins.
[6
marks]
