Six protein pairs, all with known 3D-structures, were used to evaluate different protein structure prediction tools. Firstly, alignments between a target sequence and a template sequence or structure were obtained by sequence alignment with QUANTA or by threading with THREADER, 123D and PHD Topits. Secondly, protein structure models were generated using MODELLER. The two protein structure assessment tools used were the root mean square deviation (RMSD) compared with the experimental target structure and the total 3D profile score. Also the accuracy of the active sites of models built in the absence and presence of ligands was investigated. Our study confirms that threading methods are able to yield more accurate models than comparative modelling in cases of low sequence identity (< 30%). However, a gap of 2 A (RMSD) exists between the theoretically best model and the models obtained by threading methods. For high sequence identities (> 30%) comparative modelling using MODELLER resulted in accurate models. Furthermore, the total 3D profile score was not always able to distinguish correct from incorrect folds when different alignment methods were used. Finally, we found it to be important to include possible ligands in the model-building process in order to prevent unrealistic filling of active site areas.