Sol-gel encapsulation of Cobalt centred nitrile hydratase

The nitrile hydratase from Pseudonocardia thermophila is one of the most stable NHases currently described. It is cobalt centred, and one strain of it has an entry in the PDB. Holz and co-workers have just published a paper entitled "Acrylamide Production using Encapsulated Nitrile Hydratase from Pseudonocardia thermophila in a Sol-gel matrix" in the Journal of Molecular Catalysis A. They show that their system is able to convert acrylonitrile to acrylamide neatly, demonstrating the advantages of their enzyme support system in terms of increasing the robustness of these still rather sensitive enzymes. Interesting to me is that their orthosilicate sol-gels boost this enzyme's stability in methanol to take as much as a70% v/v mix happily.

New patent from Mitsui

A Google alert has led me to a new US patent from Mitsui Chemicals Inc.

The abstract is very abstract: "A nitrile hydratase variant comprising substitution of at least one amino acid with another amino acid to improve two or more properties of nitrile hydratase by substitution of one or more and three or less amino acids".

The objective of the work under the patent is to produce a NHase variant with improved stability and high initial reaction rate.

It appears to be about Pseudonocardia thermophila variants, but I havent read all 77 pages of it since it tends to dwell on the very many changes to the sequence that they are protecting, both at the amino acid and nucleotide level.

How many clearly different structures of NHase are reported as pdb files?

Another helpful thing that the Iterative Magic Fit function within DeepView can do is provide a numerical value (based on a RMS calculation) for the similarity of the three dimensionality of the enzymes being overlaid. After concentrating on enzymes which are clearly NHases, discarding structures which are clearly mutants of an existing structure (which is either obvious from the title or the combination of author and date of submission), you get left with 13 distinct pdb files [1AHJ, 1IRE, 1UGP, 1V29, 2AHJ, 2CYZ, 2CZ6, 2D0Q, 2DPP, 2QDY, 2ZPB, 3A8O and 3HHT]. I then IMFed every one of these against each other to get a matrix of values for similarity. Basically if they had a value of less than 0.5, they look pretty much the same, greater than 1 than they look noticeably different.
The results of this showed that ALL the iron centred enzymes have RMS of <0.5 with each other which is not entirely surprising since they are all from Rhodococcus species. They all have RMS of greater than 1 for all the cobalt centred enzymes. There are two distinctly different cobalt centred arrangements: the Pseudonocardia thermophila pair of 1IRE and IUGP, and the trio of Bacillus structures of 1V29, 2DPP and 3HHT. Within these groupings they have RMS of less than 0.5, and between the groupings RMS of greater than 1.
Effectively this says to me that there are three basic structures for NHase known currently- one iron based and two cobalt based. Below is an excerpt from a table holding all the results from these calculations which shows this clustering.

PDB ID	1AHJ	1IRE
1AHJ		1.06
1IRE	1.06
1UGP	1.06	0.07
1V29	1.19	1.14
2AHJ	0.45	1.09
2CYZ	0.44	1.07
2CZ6	0.47	1.09
2D0Q	0.45	1.09
2DPP	1.22	1.16
2QDY	0.46	1.09
2ZPB	0.44	1.08
3A8O	0.47	1.11
3HHT	1.22	1.15