## 2 Results and Discussion  
### Quality Check for Docking  
It is obvious that it is important for our approach that docking of two truly interacting proteins really lead to results that are close to the native structure of the complex. As a test case we used the Barnase-Barstar complex (PDBids: Barnase: 1 RGH B, Barstar: 1A19 B, complex: 1 AY7 A:B). Barnase excreted from the bacterium _Bacillus amyloliquefaciens_ is a protein of 110 amino acids that possesses ribonuclease activity. It forms a tight complex with its inhibitor Barstar. The above input structures were used in a standard HADDOCK [8] run that resulted in 200 final complex structures, which then were analyzed in view of a correlation between RMSD to the native structure and the score calculated by HADDOCK. A short description of the HADDOCK docking algorithm can be found in the methods section.  
**Figure 1.** On the top the backbone of the native Barnase-Barstar complex structure (red) is compared to four near native solutions obtained by docking. The graph on the bottom shows the correlation between RMSD to the native structure and HADDOCK score for all 200 docking results. Pair-wise RMSD calculations are based on the coordinates of the C\(\alpha\) atoms. A description of the score calculation can be found in the experimental section. The horizontal red line separates the 10 best structures according to the calculated scores.  
In the first attempt we did not use the facility of HADDOCK to define the interacting residues since we were interested in investigating whether the correct orientation of the molecules can be found by the algorithm without additional data. Unfortunately, this approach was unsuccessful and thus we did a second test with slightly more information where one side of the interface was defined: here for Barnase the interacting residues were incorporated as ambiguous interaction restraints (these are residues number 27, 59, 60, 83, 87 and 102) and for Barstar no interface information was included. One thousand complex structures were obtained by rigid body docking and 200 of these were further refined by semi-flexible simulated annealing in torsion angle space.  
All other parameters were set to default values. This time there were several near-native structures among the 10 top ranked docking results. Ranking was based on the HADDOCK score calculation. On the top part of Figure 1 the backbones of four selected structures obtained by docking are overlaid to the native structure. It is obvious that this time near native structures could be obtained.  
This shows that it is possible to get the right conformation of a protein-protein complex even if only one interaction side is defined. However, the challenge remains to find the correct solutions among the proposed results.  
Figure 2: The two graphs show the correlation between RMSD to the native structure and Haddock score for all 200 docking results for the \(\alpha\)- Chymotrypsin - Eglin C and _Bovine_ trypsin - CMTI - 1 squash inhibitor test cases. Pair-wise RMSD calculations are based on the coordinates of the C atoms. A description of the score calculation can be found in the experimental section. The horizontal red lines separate the 10 best structures according to the calculated scores.