Hi,
We have constructed homology models based on templates of low sequence identity (27-28%). It was an enzyme and we also did some ligand-enzyme interaction study using Modeller. We did some superpositions of template and target structure and the deviation was below 1 Angstrom. We have submitted this to a journal and got a few comments. I need your advise in this regard. As you will notice that these are related to deducing and improving the quality of a homology model, I would be grateful if I can get a few tips in order to further improve the model quality in light of the listed parameters. A few of the points are concept based i.e. sequence identity and homology. Kindly comment on those. Here is the summary:
RMSD related: The backbone could deviate from the true structure by a few angstroms and the impact of such deviation for enzyme catalysis could be huge. Therefore, the application of such models is often limited. Trying to deduce the specific interactions between the active site residues and a substrate based on such a model is thus questionable. The rmsd between the model and the template was ~1 angstrom is not the evidence of high quality of the models. The term rmsd is being mis-understood. In structure prediction, the rmsd between a homology model of the target protein and the X-ray structure of the same target protein was often calculated as the indication of the quality of the models, but not the rmsd between the model and the X-ray structure of the template.
Sequence Related: The multiple alignment was only used for the phylogenetic analysis, whereas the pairwise alignments were used as a starting point to create the models. This procedure is conceptually wrong, since the alignment created by PSI-BLAST is not necessarily the best one. In fact, PSI-BLAST makes a pairwise alignment to find the most similar sequences, not to find the best alignment between the sequences. Moreover, with such a low sequence similarity (less than 30%) the best procedure to be sure of a good starting alignment is to perform a multiple alignment, and in case to use also predictions of the position of secondary structure elements, etc in order to improve as much as possible the quality of the alignment (see refs. to the CASP competitions). Then, the alignment between template(s) and model should be extrapolated by the multiple sequence alignment.
Homolgy Definition: The term "homology" simply indicates the presence of a common evolutionary origin between two biological entities: therefore, two proteins are homologous or not. Somebody said that talking about "X% of homology" would be more or less the same as talking about a women who is "X% pregnant". Instead, it is correct to say that two proteins have X% of their amino acids identical.
Energy Minimization: The authors do not have the energy minimized structure, which is a must for carrying out interaction analyses and if not done, leads to incorrect interpretations. They have assumed the program Modeller provides a proper energy-minimized structure with 'automodel' environment. However, published literature that use Modeller in such an environment still have been found to energy minimize the structure and only then use further.
Thank you asmat