Nitrile hydratases (EC) are bacterial enzymes that catalyse the hydration of nitrile compounds to the corresponding amides. They are used as. Characterization of a Nitrilase and a Nitrile Hydratase from Pseudomonas sp. Strain UW4 That Converts IndoleAcetonitrile to IndoleAcetic Acid. Nitrile hydratase (NHase) was discovered in our laboratory. This enzyme was purified and characterized from various microorganisms. NHases are roughly.
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Some pathways intersect at a common intermediate, implying that that route may be the predominant one.
Unraveling the Catalytic Mechanism of Nitrile Hydratases
The advantage of having a complex multiroute biosynthetic system protects the bacterium against the loss of any particular pathway, since IAA production can be fluxed through an alternate circuit The genome of an IAA-producing, plant growth-promoting bacterium, Pseudomonas sp.
UW4, has recently been completely sequenced Nitrile hydratase genes that encode enzymes potentially involved in the biosynthesis of IAA were identified These genes appear to be part of at least two different IAA biosynthesis pathways.
Thus, the present study is directed toward deciphering the IAA biosynthesis pathways operating nitrile hydratase Pseudomonas sp. UW4 by isolating, purifying, and characterizing two of the implicated enzymes.
We hypothesize that in the former pathway, the tryptophan precursor is first converted to the IAOx intermediate by an nitrile hydratase enzyme. The focus of the work reported here is the isolation, purification, and partial characterization of the Nit and NthAB enzymes.
Examination of a plot of log kcat versus pH revealed a nitrile hydratase curve that yielded a pKES1 value of 5.
Nitrile hydratase - Wikipedia
The slopes of the asymptotes, calculated as described previously 30of the acidic and basic limbs of log kcat versus pH for PtNHase are 1, indicating that one group is ionized on each limb. These data indicate nitrile hydratase one ionizable group pKES1 must be deprotonated and that a second ionizable group must be in the protonated form nitrile hydratase in the ES complex for catalysis to occur.
Assignment of the observed pKES values is difficult, but likely candidates for pKES1 are the deprotonation of the metal-coordinated sulfinic acid putative pKa in NHase of 7. A sequence nitrile hydratase genome of the choanoflagellate Monosiga brevicollis was suggested to encode for a nitrile hydratase.
Similar nitrile hydratase genes consisting of nitrile hydratase fusion of the beta and alpha subunits have since been identified in several eukaryotic supergroups, suggesting that such nitrile hydratases were present in the last common ancestor of all eukaryotes.
Thiocyanate hydrolase SCNase is a cobalt-containing metalloenzyme with a cysteine-sulphinic acid ligand that hydrolyses thiocyanate to carbonyl sulphide and ammonia [ PMID: The two enzymes, nitrile hydratase and SCNase, are homologous over regions corresponding to almost the entire coding regions of the genes: It excludes the thiocyanate hydrolase gamma subunit of Thiobacillus thioparus, a sequence that appears to have evolved from within the family of nitrile hydratase alpha subunits nitrile hydratase which differs by several indels and a more rapid accumulation of point mutations.
Novel non-heme iron center of nitrile hydratase with a claw setting nitrile hydratase oxygen atoms.