My presentation focused on a method to predict protein flexibility – PredyFlexy. There is a webserver, and it is described in their paper (de Brevern et. al 2012). The method is covered much more explicitly in the paper Predicting protein flexibility through the prediction of local structures (Bornot et al. 2011). This work builds on earlier papers (which are mentioned on the front page of the webserver, some of which I will mention later). In terms of people, Alexandre G. de Brevern, Aurelie Bornot, and Catherine Etchebest are authors common to all of these papers.
The concept is simple; there is a library of ‘Local Structure Prototypes’ or LSPs. These LSPs are 11 residue fragments, and have structure and flexibility associated with them, which are derived from a set of protein structures and molecular dynamics simulations. Each LSP has a SVM (support vector machine) ‘expert’ to score how likely a given sequence profile is to have said structure. A 21 residue window is used to determine the LSP of the central 11 residues within this window.
So, the user inputs a sequence, which gets Psi-Blasted to give a sequence profile. The 5 most probable LSPs for each atom are determined using the SVM experts. Then the predicted flexibility of each atom is given by the average flexibility of these 5 LSPs. There is a confidence index to the predictions, which comes from assessing the discriminative power of the SVMs. Regions predicted to have LSP with more accurate SVMs will have a high confidence index.
So, the concept seems simple, but you are probably wondering, what are these LSPs? To answer that, we have to delve into the literature. ‘Bayesian probabilistic approach for predicting backbone structures in terms of protein blocks‘, published in 2000, and with Alexandre G. de Brevern as first author, is the sensible place to start. This introduced the concept of protein blocks, which is in effect a structural alphabet. These are 5 residue fragments (described by 8 dihedral angles), and there are 16 of them (there are pretty pictures in the supplementary data 1 of this paper). Local structure prototypes are made up of these protein blocks, which were first used to predict structure, in a 2006 paper – Assessing a Novel Approach for Predicting Local 3D Protein Structures from Sequence (again, with the same authors). LSPs are 11 residue fragments, made from 7 overlapping protein blocks. Obviously there are lots of combinations to the protein blocks that can give 11 residue fragments. These combinations are clustered into 120 groups. Each group is represented by the fragment within the cluster that is closest to all other fragments within the cluster, based on C-alpha RMSD. Hence 120 LSPs.
The other pertinent question is, where does the flexibility data come from? Well, they did some MD simulations (see Bornot et al. 2011 for details) and took b-factors from the structures. They normalised these, by calculating the number of standard deviations away from the mean for each structure. Each LSP was attributed a b-factor and RMSF, which was the mean value for the central residue of every instance of the LSP in the data set. Additionally, each fragment in the data set was classed as ‘rigid’, ‘flexible’, or ‘intermediate’ based on its normalized RMSF and B-factor. Each LSP was given a probability of belonging to each of these classes based on the frequency of fragments that belonged to that LSP being in each class. The figure here (taken from Bornot et al. 2011) shows the interesting weak correlation between normalised B-factor and normalised RMSF.
Bornot et al. 2011 also gives us a guide to the ability of this prediction method (see table II). In predicting the class of fragments (rigid, intermediate or flexible), it gets the correct class about half the time. For 40% of rigid and flexible cases, the class is predicted as ‘intermediate’. Prediction rate is also strongly correlated with flexibility – more flexible regions have much poorer prediction rates. Which is not great, as we already know that most alpha helices are rigid. However, the confidence index does give a good guide as to what to trust. I could speculate that might results in an output that tells us that helices and sheets are definitely rigid, and other elements are possibly flexible. Which would not be particularly useful, but given there are few comparable tools, something is better than nothing.
Protein flexibility is hard; experimentally determining it is difficult (and even MD simulations take a while), and people can argue about how relevant the experimental methods are (and we frequently do in our group meetings). So, like most predictive methods, a relatively fast (and simple) way to get some information about your problem is always going to be useful. If only to guide you to where you might focus your attention.