Thesis Title: Subtitle - NMR Spectroscopy Research Group

10 Chapter 1. Introduction.
Protein-ligand interaction: John et al. (John et al., 2006) showed how PCS restraints can
be used to determine the structure of protein-ligand complexes. The approach is of major interest
for drug screening. In a first step, the Δχ-tensor is determined for the target protein. This is the most
complicated task because the protein can be large, making the assignment of chemical shifts
difficult. However, in the context of drug screening this step needs to be performed only once. Each
ligand that is being screened is isotopically labeled, and a paramagnetic spectrum of the ligand in
complex with the protein is recorded. The assignment of a ligand spectrum and the corresponding
Δχ-tensor are swiftly and easily obtained. The two Δχ-tensors from ligand and protein being the
same by definition, a simple superimposition of them leads to the rigid body structure of the
complex. A molecular dynamics package is further used to locally refine the conformation of the
protein on the contact surface. John and coworkers demonstrated the approach with the thymidine
nucleotide as the ligand binding to the ε subunit of DNA polymerase III. The determined thymidine
structure was found to have a very similar binding mode compared to the thymidine
monophosphate present in the reference crystal structure.
Protein-protein interaction: Pintacuda et al. (Pintacuda et al., 2006) described a protocol
to solve the structure of a protein-protein complex using only PCSs and illustrated the protocol on
the example of the N-terminal domain of the subunit ε and subunit θ of the E. coli DNA
polymerase III. Again the method relies on the determination of the Δχ-tensors, first relative to one
molecule (ε, Figure 1.8.a) and then relative to the second molecule (θ, Figure 1.8.b), followed by
the superimposition of the Δχ-tensor frames (Figure 1.8.c). Such an approach is particularly
relevant considering the difficulty to co-crystallize proteins in complexes, compared to the
crystallization of the components separately. An alternative has been to use RDCs (McCoy et al.,
2002), but the relative orientation and location of the two rigid bodies obtained with PCSs is more
accurate compared to what would result from a complex build from RDC data, since PCSs yield
simultaneously orientation and distance information, while RDC data lack distance information and
are sensitive to small fluctuations of NH bond orientation. The resulting rigid-body docked
complex could exhibit sterical clashes that would need to be resolved with a molecular refinement
package. The final result is valuable as input for docking refinement software considering that the
most difficult part of docking two molecules in a complex is to obtain with confidence the
approximate binding sites.