Proper NCBI numbers for August

I have been looking for a number for the actual number of proper NHase alpha subunit sequences on record. I have done this by really combing through the NCBI protein records using a BLASTp approach for similarity, and excluding sequences that aren't an appropriate length nor have the usual active site binding motif. I have ignored environmental samples, and dropped out data from PDB files.
As of the start of this week (8th Aug 2011) I reckon there are about 190 NHase alpha sequences. That breaks down as about 25% iron and 75% cobalt centred. There are four from eukaryotic organisms (three marine and one plant), and the rest are... not.

Nitrile hydratases and the Gordon and Betty Moore Foundation’s Marine Microbiology Initiative

Since the most exciting new source of nitrile hydratase sequences has been marine organisms recently, I thought I would have a quick look at the preponderance of nitrile hydratase sequences in marine bacteria. I decided to look at the Moore Foundation's Marine Microbiology Initiative which was run by JCVI. I used the alpha sequence from the nitrile hydratase from Rhodopseudomonas palustris CGA009 as a probe sequence and BLASTped this sequence against the genomes produced by this initiative. This was a slightly more tedious job than I imagined because the BLAST server at JCVI wasn't working, so I had to work through a NCBI BLASTp input, which involves a fair bit of cutting and pasting, AND the taxonomy of the names listed isn't always the same in both databases. Anyway, of the 177 marine microbes which had their genomes listed, 30 produced sequences which had query coverages of greater than 68%, and all were auto-annotated as nitrile hydratase alpha subunits and contained the appropriate metal binding sequence.

Hyperthermophilic nitrile hydratases?

There are plenty of reports of nitrilases which have been cloned from organisms with names that denote obviously heat-loving tendencies- an example is the “Cloning, overexpression, and characterization of a thermoactive nitrilase from the hyperthermophilic archaeon Pyrococcus abyssi” by Mueller, Egorova, Vorgias, Boutou, Trauthwein, Verseck and Antranikian in Protein Expr Purif. 2006, 47(2), 672-81. It is quite noticeable that NHases do not appear to have the same hyperthermophilicity. If you search the translated genomes of the four organisms which produce nitrilases in this paper (Pyrococcus abyssi, Pyrococcus horikoshii, Pyrococcus furiosus, and Aeropyrum pernix) by using BLASTp against the alpha subunit of my favourite cobalt centred NHase CGA009 or the “local” iron centred NHase AJ270, you get just about nothing that shows any similarity.

Something that does stand out from examination of all the current PDB files of NHases that almost everyone says their NHase is thermostable or thermophilic. I get the impression that this labelling of these generally sensitive enzymes in itself tells a story.