Transcript Representations of 3D Structures

A real example.
The rat fatty acid acyl carrier
protein. Involved in fatty acid
biosynthesis and part of a
larger subunit, the synthase,
Is it structured by itself??
Summary of the Sequential and Secondary NOEs observed for
rat FAS ACP - most definitely structured
NHi-NHi+1
iNHi+1
iNHi+1
GDGEAQRDLVKAVAHILGIRDLAGINLDSSLADLGLDSLMGVEVR
D
D
D
DD
D
D
D
NHi-NHi+2
Hi-NHi+2
Hi-NHi+3
Hi-NHi+4
Hi-Hi+3
CSI
J
0-00000---------+-0-0--0+--+0+---+00+-0-----+ ++ -------+--+++++ +++-+++ --- ----- --QILEREHDLVLPIREVRQLTLRKLQEMSSKAGSDTELAAPKSKN
NHi-NHi+1
iNHi+1
iNHi+1
D
D
D
D
DD D
D
D DD
NHi-NHi+2
Hi-NHi+2
Hi-NHi+3
Hi-NHi+4
Hi-Hi+3
CSI
J
-----0+-+0++--0--00+--------00000000+0+00-00
-++-+++ - -- -+-+ - -+- +++++++
+++
What next? STRUCTURE CALCULATIONS
•From NOE I know close atom-atom distances, but that doesn’t give a structure
•The information you have up to this stage is a list of distance constraints
•The structure can be determined by inputting this information to computer
minimization software.
•The computer program also contains information about amino acids, bond
lengths/angles and standard information about atom-atom interactions such as
minimum distance (i.e. Van der Waals radii)
•With all this information you can generate a model of the structure.
Important: NMR gives you a number of possible solutions
(all almost identical, rmsd <1Å), This can range from 5-20 models
X-ray crystallography give one average structure
NMR structures can be averaged to give one average structure as well
A simulated annealing trajectory over the first few picoseconds
4 helices begin to
‘condense’
Unfolded
Correctly folded
Representations of 3D
Structures
C
N
Precision is not Accuracy
These 2D methods work for proteins up to about 100 amino acids,
and even here, anything from 50-100 amino acids is difficult.
We need to reduce the complexity of these 2D spectra.
1
16
1
H
R2
O
HN
16
O
12
C
12
C
14
N
14
N
12
C
12
C
1
R1
1
H
HN
We can start by
replacing 14N with
15N, a spin 1/2
nucleus.
HSQC of rat FAS ACP
15N shift of nitrogen of amide bond
1H-15N
H
N
1H
Chemical Shift
X
89!