How have the database numbers changed in a year?

Last year I did a quick text search for "nitrile hydratase" as a search term under proteins on the NCBI website. This gave me 2869, of which 1042 were RefSeq data. Today when I checked there are 3573 (+25%), of which 1369 (+31%) were RefSeq.
No new PDB files have been deposited of nitrile hydratases since March 2011 which was the NHase from Pseudomonas putida.

Comparing the access of the active site of 3QXE and 3HHT

If you orient the pdb file of the Geobacillus which is coded as 3HHT as I have done for 3QXE and 2QDY in the previous posts, you get the following two images.

The cobalt is no longer visible. However if you twirl the structure around you can find a view of it through a different aperture. I wonder how this impacts on the selectivity of this enzyme.

Looking inside for the active site of 3QXE

This is just a couple of rough pictures of 3QXE (the nitrile hydratase from Pseudomonas putida), once again from PyMol, showing a ribbon view of it with the active site cobalt-binding residues shown as CPK spheres, followed by a surface model with some of the active site just visible inside. I have been careful to keep it oriented similarly to the AJ270 nitrile hydratase in the previous post.

Salt bridges and hydrogen bonding

I have been playing with PISA tool as it offered insight into size and nature of the interface between the two subunits in NHases. I stuck in the PDB data for the most distinct cobalt-centred NHases- 1UGP, 1V29 and 3HHT, and PISA suggested that these have 61, 67 and 65 hydrogen bonds (respectively) between the alpha and beta subunits, and between 26 and 29 salt bridges across this interface. Compare these numbers with that for 2QDY (the iron-centred NHase from AJ270)... 51 hydrogen bonds and 8 salt bridges. That looks quite a difference... sadly things might not be so clear-cut in grouping these enzymes by metal centre- the newest PDB for 3QXE gives an average of 53 hydrogen bonds and 15 salt bridges, and that is cobalt-centred. Furthermore I do wonder, with such a small set of structures, whether we are seeing a distinction based on the fact that the first three cobalt-centred PDBs are of enzymes specifically tagged as "thermophilic", and AJ270 and Pseudomonas putida aren't. [I also looked at 2DPP and got 62 hydrogen bonds and 27 salt bridges].

Cobalt binding in 3QXE

Above is an excerpt from Figure 6C in "Evidence for Participation of Remote Residues...etc by Ondrechen and Ringe which very nicely shows how the cobalt ion is bound into a nitrile hydratase.  There are three sulphur based points of attachment- aC112, aC115 and aC117, the latter two as sulfinic acids and two amide nitrogen points of attachment, aS116 and aC117. It is really clearly shown in this view how the arginine bR52 preorganizes the sulfinic cysteines and amide nitrogens into a planar SSNN arrangement.

The paper that goes with 3QXE

The paper which describes the work that lead to the pdb file 3QXE and its mutant brethren has just been put up as an accepted manuscript for the ACS journal, Biochemistry. Its corresponding authors are Mary Jo Ondrechen and Dagmar Ringe, and it goes by the inviting title of "Evidence for Participation of Remote Residues in the catalytic activity of Co-type Nitrile hydratase from Pseudomonas putida". They have been looking at which residues in the outer shells around the active site influence activity.

PDBs on a phylogenetic tree

There is a figure of a phylogenetic tree in the Prasad Balla review from 2010 which gives a separation of the different currently known sequences. I have very roughly stitched the tree together from the two parts given on two pages of the review and then I have dropped on top the rough positions of the three most distinct cobalt centred PDB files (1V29, 1UGP and 3QXE) and one of the rhodococcus iron centred files (1AHJ). It is interesting that workers seem to be producing PDB files solely from the right hand half of this tree.

3QXE- the new NHase pdb file- from Pseudomonas putida

I thought I'd have a look to see how different the new pdb file was from those already in the public domain.
First of all a ClustalW alignment of the alpha chains shows that it has a score of 52 with 1V29 and a score of 54 with 1UGP. (For reference, 1V29 and 1UGP have a score of 59 between them). The numbers might not be completely robust because I note that the modified cysteines are not dealt with the same in the three amino acid sequences. The central portions of the sequence are very similar, with most of the diversity at the ends (see below: 1V29 then 1UGP and finally 3QXE)

Using DeepView's RMS measure of 3D similarity after an Iterative Magic Fit of  the new pdb with the files for 1V29 and 1UGP enzymes you get two numbers of approximately 1-1.05 (for reference between 1V29 and 1UGP the value is 1.15). This all suggests that this new file describes a new distinct structural arrangement.