Analysis: Results from thermal stability shift and competition binding assays correlate well

Several large kinase inhibitor screens have been published in recent years. Two of the largest come from Stefan Knapp’s lab and Ambit, respectively. The former group used a temperature shift assay to measure the change in thermal stability of 60 human serine/threonine kinases that is caused by the binding of each of 156 kinase inhibitors (Fedorov et al., 2007). The latter group used a competition a competition binding assay to measure the dissociation constants (Kd) for 38 kinase inhibitors and 290 distinct kinases (Karaman et al., 2008).

The two screens are not directly comparable because one measures temperature shifts whereas the other measures dissociation constants. To see if it possible to convert temperature shift values to Kd values, I asked Damian Szklarczyk (who is a Ph.D. student in my group) to map all data from both screens onto a common set of chemical and protein identifiers, extract all inhibitor-kinase pairs that were measured in both assays, and make a scatter plot of -log(Kd) as function of temperature shift. The result was a set of 704 pairs of temperature shift and Kd values. In the plot below, inhibitor-kinase pairs for which binding was not observed in the competition binding assay were defined to have a Kd of 10 microM, and negative values from the temperature shift assay were treated as zero temperature shift.

The plot shows that the two assays are in very good agreement, which is surprising considering that the assays are fundamentally very different and were run using different expression constructs for several of the kinases. The linear Pearson correlation coefficient is 0.92 when excluding the one obvious outlier shown in red (BIRB796 vs. MAPK11; this appears to be a false negative in the competition binding assay).

The linear fit gives an intercept with the y-axis of 4.9223, which implies that a temperature shift of zero (i.e. no binding according to the temperature shift assay) does not translate precisely into a Kd of 10 microM (i.e. no binding according to the competition binding assay). We thus did a second linear regression in which we forced the intercept with the y-axis to 5 (red regression line in the plot). We thereby at the calibration function -log(Kd) = 5+0.244*Ts, which allows us to to convert temperature shifts to Kd values. We have thereby managed to put the measurements from the two kinase inhibitor screens onto a common basis that facilitates direct comparison and integration.

Full disclosure: I have an on-going collaboration with Stefan Knapp’s lab related to screening of kinase inhibitor.

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Resource: Second Life Interactive Dendrogram Rezzer (SLIDR)

About half a year ago, I began experimenting with Second Life as a tool for virtual conferences (I should add that my experiences have since improved). However, I believe that imitating real life in a virtual world is not necessarily the best way to use the technology – it may be better to use virtual reality for doing the things that are difficult to do in the real world. A good example of this is Hiro’s Molecule Rezzer, which is one of the best known scientific tools in Second Life. It, and its much improved successor Orac, allows people to easily construct molecular models of small molecules in Second Life.

After speaking with several other researchers in Second Life, who like I are interested in evolution, I set out to build a similar tool for visualization of phylogenetic trees. The result is SLIDR (Second Life Interactive Dendrogram Rezzer), which based on a tree in Newick format constructs a dendrogram object. The first version of SLIDR can handle trees both with and without branch lengths; however, I have not yet implemented support for labels on internal nodes or for bootstrap values.

The picture below shows an example of a dendrogram that was automatically generated by SLIDR based on a Newick tree:

There is a bit more to SLIDR than this, though. After the dendrogram has been built, it can be loaded with a photo and/or a sound for each of the leaf nodes. When click on a node, the corresponding sound will be played and the photo will be shown on the associated screen (the white box in front of which I stand):

I plan to work with collaborators in Second Life to construct dendrograms for evolution of bats (including their echolocation sounds and photos of the animals) and for the fully sequenced Drosophila genomes. Please do hesitate to contact me if you would like to use SLIDR on another project. I intend to make SLIDR available as open source software once I have implemented support for the full Newick format.

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