Transcript Document

Basic protein structure and
stability V:
Even more protein anatomy
Biochem 565, Fall 2008
09/05/08
Cordes
Tertiary structure in proteins
Tertiary is the level of protein structural hierarchy above
secondary, and involves:
• The number and order of secondary structures in the
sequence (connectivity) and their arrangement in
space. This defines a protein’s tertiary fold (more
on this later) also called its global topology
• Pattern of contacts between side chains/backbone,
including and especially contacts between residues in
different regions of the sequence (long-range)
• Outer surface and interior--what’s inside/outside
• Limited to interactions within a single polypeptide
chain--interaction between chains is quaternary
Supersecondary structures/structural motifs
•
•
•
just as there are certain secondary structure elements that are common, there
are also particular arrangements of multiple secondary structure elements that
are common
note that I don’t consider a beta-sheet a secondary structure element,
because it is not regular and contiguous in the sequence--I consider the
individual strands to be secondary structure elements, while the sheets
formed from these strands are supersecondary structures (really an aspect of
tertiary structure)
supersecondary structures emphasize issue of topology in protein structure
b-a-b motif
greek key motif
Topology: differences in connectivity
a four-stranded antiparallel b sheet can
have many different topologies based on the order
in which the four b strands are connected:
• example:
“up-and-down”
“greek key”
Topology: differences in handedness
•
•
example: An extremely common supersecondary structure in proteins
is the beta-alpha-beta motif, in which two adjacent beta-strands are
arranged in parallel and are separated in the sequence by a helix which
packs against them.
if the two parallel strands are oriented to face toward you, the helix can
be either above or below the plane of the strands.
huge preference for right-handed
arrangement in proteins
Topology/geometry: beta-sheet
twisting
Beta-sheets are not flat:
they have a right or left
handed “twist”, and
essentially all beta-sheets
in proteins have a right-handed
twist like that seen in
flavodoxin (1czn) at right.
See Richardson article for naming
convention, and ideas about the
origin of a preference for the
right-handed twist.
Visualizing topology--TOPS cartoons
Cu/Zn superoxide dismutase
all anti-parallel
beta structure
sheet 1
sheet 2
this is a TOPS
cartoon of the
structure at left
• lines = loops
• up triangles = up-facing b strands
• if loop enters from top, line drawn to • down triangles = down-facing b strands
center
• horizontal rows of triangles = b sheets
• if loop enters from bottom, line
(beta barrel would be a ring of triangles)
drawn to boundary
• circles = helices
Contact maps of protein structures
-both axes are the sequence of the protein
map of Ca-Ca distances < 6 Å
near diagonal: local
contacts in the
sequence
off-diagonal: long-range
(nonlocal) contacts
rainbow ribbon diagram
blue to red: N to C
1avg--structure of triabin
Contact maps of protein structures
-both axes are the sequence of the protein
map of Ca-Ca distances < 6 Å
rainbow ribbon diagram
blue to red: N to C
Structure of n15 Cro
Contact maps of protein structures
-both axes are the sequence of the protein
map of all heavy atom distances
< 6 Å (includes side chains)
rainbow ribbon diagram
blue to red: N to C
Structure of n15 Cro
Surface and interior of globular
proteins
solvent accessible surface
molecular surface
residue fractional accessibility
pockets and cavities
“hydrophobic core”
ordered waters in protein structures
“Accessible Surface”
represent atoms as spheres w/appropriate
radii and eliminate overlapping parts...
mathematically roll a
sphere all around that
surface...
the sphere’s
center traces
out a surface
as it rolls...
Lee & Richards, 1971
Shrake & Rupley, 1973
Now look at a cross-section (slice) of a protein structure:
Inner surfaces here are van der Waals. Outer surface is that traced out by the
center of the sphere as it rolls around the van der Waals’ surface. If any part of
the arc around a given atom is traced out, that atom is accessible to solvent.
The solvent accessible surface of the atom is defined as the sum the arcs
traced around an atom.
there’s not much solvent accessible surface
in the middle
van der Waals
surface
solvent
accessible
surface
from
Lee &
Richards,
1971
arc traced around atom
“Accessible surface”/“Molecular surface”
note: these are alternative ways of representing the same reality:
the surface which is essentially in contact with solvent
• molecular and accessible surfaces are both useful
representations, but molecular surface is more
closely related to the actual atomic surfaces. This
makes it somewhat better for visualizing the texture
of the outer surface, as well as for assessing the
shape and volume of any internal cavities.
• you will hear the term Connolly surface used often,
after Michael Connolly. A Connolly surface is a
particular way of calculating the molecular surface.
The accessible surface is also occasionally called the
Richards surface, after Fred Richards.
Molecular surface of proteins
depiction of heavy atoms (O,
N,C, S) in a protein as van der
Waals spheres
depiction of the corresponding
“molecular surface”--volume contained
by this surface is vdW volume plus
“interstitial volume”--spaces in between
The irregular surface of proteins:
pockets and cavities
•
a pocket is an empty
concavity on a protein
surface which is
accessible to solvent
from the outside.
•
a cavity or void in a
protein is a pocket
which has no opening
to the outside. It is
an interior empty
space inside the
protein.
Pockets and cavities can be critical features of proteins in terms of
their binding behavior, and identifying them is usually a first step in
structure-based ligand design etc.
Fractional accessibility
•
•
•
•
calculate total solvent accessible surface of protein structure (also can
calculate solvent accessible surface for individual residues/sidechains
within the protein)
can also model the accessible surface area in a disordered or unfolded
protein using accessible surface area calculations on model tripeptides
such as Ala-X-Ala or Gly-X-Gly.
from these we can calculate what fraction of the surface is buried
(inaccessible to solvent) by virtue of being within the folded, native
structure of the protein.
this is done by dividing the accessible surface area in the native
protein structure by the accessible surface in the modelled
unfolded protein. That’s the fractional accessibility. The residue
fractional accessibility and side chain fractional accessibility refer to the
same thing calculated for individual residues/sidechains within the
structure.
Accessible surface area in globular protein structures
Accessible surface area As in native states of proteins is a non-linear
function of molecular weight (Miller, Janin, Lesk & Chothia, 1987):
As = 6.3Mr0.73
`
where Mr is molecular wt
This is an empirical
correlation but it comes
close to the expected
two-thirds power law
relating surface area to
volume or mass for a set
of bodies of similar shape
and density.
How much surface area is buried when a protein
adopts its native structure in solution?
•
estimate total accessible surface area in extended/disorded polypeptide
chain using the accessible surface areas in Gly-X-Gly or Ala-X-Ala
models. This is a linear function of molecular weight
At = 1.48Mr + 21
•
the total fractional accessibility is As/At ,and the fraction of surface area
buried is 1-
•
As /At
What is the total fractional surface area buried for a protein of
molecular weight 10,000? 20,000? Is the fraction higher for small
proteins or large?
Distribution of residue fractional accessibilities
note that a sizeable group are completely buried
(hatched) or nearly completely buried
note broad distribution among non-buried
residues, and mean fractional
accessibility for non-buried residues
of around 0.5
note that few residues are
completely exposed to
solvent, but that fractional
accessibility of >1 is possible
from Miller et al,
1987
Buried residues in proteins
•the fraction of buried residues (defined by 0% or 5%
ASA cutoffs) increases as a function of molecular
weight--for your average protein around 25% of the
residues will be buried. These form the core.
size class
mean Mr
small
medium
large
XL
all
8000
16000
25000
34000
fraction of buried residues
0% ASA
5% ASA
0.070
0.154
0.107
0.240
0.139
0.309
0.155
0.324
0.118
0.257
Residue fractional accessibility correlates with free
energies of transfer for amino acids between water
and organic solvents
• (Miller, Janin, Lesk & Chothia, 1987)
• (Fauchere & Pliska, 1983)
• the interior of a protein is akin to a
nonpolar solvent in which the nonpolar
sidechains are buried. Polar sidechains,
on the other hand, are usually on the
surface. However, some polar side chains
do get buried, and it must also be
remembered that the backbone for every
residue is polar, including those with
nonpolar side chains. So a lot of polar
moieties do get buried in proteins.
The hydrophobic core of a small protein:
N15 Cro
0% ASA:
Pro 3
Leu 6
Ala 16
Val 27
Ile 36
Ile 44
< 5 % ASA:
Met 1
Ala 17
Val 20
note that some polar residues
Gln 41
are buried
Ser 54
11 of 66 ordered residues have less than 5% ASA
The outer surface: water in protein structures
Structures of water-soluble
proteins determined at
reasonably high resolution
will be decorated on their
outer surfaces with water
molecules (cyan balls) with
relatively well-defined
positions, and waters may
also occur internally
Water is not just surrounding
the protein--it is interacting
with it
Water interacts with protein surfaces
Most waters visible in crystal structures make hydrogen bonds
to each other and/or to the protein, as donor/acceptor/both
second shell water:
only contacts other waters
first shell waters:
in contact with/
hydrogen bound
to protein