Gene Ontology annotation through association of InterPro records with GO terms
Use of the ND evidence code for Gene Ontology (GO) terms
Electronic Gene Ontology annotations created by ARBA machine learning models
Crystal structure of the worm NitFhit Rosetta Stone protein reveals a Nit tetramer binding two Fhit dimers.
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Nit-domain proteins of the nitrilase superfamily have a presumptive Cys-Glu-Lys catalytic triad; this is a structure paper on C. elegans NitFhit and does not demonstrate any catalytic substrate.
"Nit monomers possess a new alpha-beta-beta-alpha sandwich fold with a presumptive Cys-Glu-Lys catalytic triad"
The nitrilase superfamily: classification, structure and function.
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Only branch 1 of the nitrilase superfamily has demonstrated nitrilase activity; most branches are amidases or amide-condensation enzymes, so family membership does not imply nitrilase activity.
"Despite historical classification of all of these sequences as nitrilase-related, only one branch is known to have nitrilase activity, whereas eight branches have apparent amidase or amide-condensation activities."
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Nitrilase-superfamily database annotations are frequently applied without supporting activity data.
"Automated sequence searching easily identifies predicted polypeptides as members of the nitrilase superfamily, but many database annotations have been applied haphazardly."
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The superfamily shares a conserved Glu-Lys-Cys catalytic triad.
"Essentially all members of the nitrilase superfamily have a conserved, apparent catalytic triad of glutamate, lysine and cysteine"
Nit1 is a metabolite repair enzyme that hydrolyzes deaminated glutathione.
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Human NIT1 and its characterized yeast ortholog are deaminated-glutathione amidases (metabolite repair), not nitrilases; the yeast enzyme studied is the gene named NIT2 (YJL126W), a distinct paralog from YIL164C.
"both the mammalian Nit1 and its yeast ortholog are amidases highly active toward deaminated glutathione (dGSH; i.e., a form of glutathione in which the free amino group has been replaced by a carbonyl group)"
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Even the closest characterized relatives are amidases whose function was long unresolved; nitrilase activity is not their function.
"the Nit1 protein does not hydrolyze efficiently α-ketoglutaramate (a known physiological substrate of Nit2), and its actual enzymatic function has so far remained a puzzle"
UniProt entry P40447 (NIT1_YEAST), Saccharomyces cerevisiae
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NIT1 belongs to the carbon-nitrogen hydrolase superfamily (nitrilase family) and PROSITE predicts a Glu/Lys/Cys catalytic triad.
"Belongs to the carbon-nitrogen hydrolase superfamily."
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UniProt cautions that NIT1 may be the product of a pseudogene and appears to be the N-terminal part of a nitrilase-like protein formed with YIL165C.
"Could be the product of a pseudogene. NIT1/YIL164C seems to be"
Bioinformatics analysis of NIT1 (catalytic triad, truncation, orthology)
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The Glu-Lys-Cys catalytic triad (E44,K135,C169) is present and correctly positioned in NIT1, aligning to the triad of human NIT1/NIT2 and Arabidopsis NIT1; NIT1 is not a triad-loss pseudoenzyme.
"All three canonical nitrilase-superfamily triad residues are present at the predicted positions."
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NIT1 (YIL164C) covers only the N-terminal ~2/3 of a nitrilase fold and YIL165C the C-terminal ~1/3; their concatenation spans a full-length domain.
"NIT1 (cols 1-270) and YIL165C (cols 271-406) tile the domain almost exactly end-to-end with no overlap"
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Both yeast ORFs fall in the uncharacterized PANTHER subfamily PTHR46044:SF1, not in any substrate-defined branch; no specific substrate can be assigned.
"No specific catalytic substrate can be assigned."
Falcon deep research report for NIT1 (YIL164C)
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The report incorrectly equates YIL164C ("scNit1") with the characterized deaminated-glutathione amidase and transfers that function, kinetics and crystal structure to YIL164C. This is a paralog conflation: the dGSH amidase is the yeast gene NIT2 (YJL126W) per Peracchi 2017 Table 1 and SGD, whose triad numbering (Glu45-Lys127-Cys169) differs from YIL164C's predicted Glu44-Lys135-Cys169.
"*S. cerevisiae* NIT1 (YIL164C, P40447) encodes a deaminated glutathione (dGSH) amidase"