NRL4_ARATH
ID NRL4_ARATH Reviewed; 355 AA.
AC P46011; Q8L9L4;
DT 01-NOV-1995, integrated into UniProtKB/Swiss-Prot.
DT 01-NOV-1995, sequence version 1.
DT 25-MAY-2022, entry version 151.
DE RecName: Full=Bifunctional nitrilase/nitrile hydratase NIT4;
DE EC=3.5.5.1;
DE EC=3.5.5.4;
DE EC=4.2.1.65;
DE AltName: Full=Cyanoalanine nitrilase;
DE AltName: Full=Nitrilase 4;
GN Name=NIT4; OrderedLocusNames=At5g22300; ORFNames=MWD9.8;
OS Arabidopsis thaliana (Mouse-ear cress).
OC Eukaryota; Viridiplantae; Streptophyta; Embryophyta; Tracheophyta;
OC Spermatophyta; Magnoliopsida; eudicotyledons; Gunneridae; Pentapetalae;
OC rosids; malvids; Brassicales; Brassicaceae; Camelineae; Arabidopsis.
OX NCBI_TaxID=3702;
RN [1]
RP NUCLEOTIDE SEQUENCE [MRNA].
RC STRAIN=cv. Landsberg erecta;
RX PubMed=8022831; DOI=10.1073/pnas.91.14.6649;
RA Bartel B., Fink G.R.;
RT "Differential regulation of an auxin-producing nitrilase gene family in
RT Arabidopsis thaliana.";
RL Proc. Natl. Acad. Sci. U.S.A. 91:6649-6653(1994).
RN [2]
RP NUCLEOTIDE SEQUENCE [LARGE SCALE GENOMIC DNA].
RC STRAIN=cv. Columbia;
RX PubMed=9501997; DOI=10.1093/dnares/4.6.401;
RA Nakamura Y., Sato S., Kaneko T., Kotani H., Asamizu E., Miyajima N.,
RA Tabata S.;
RT "Structural analysis of Arabidopsis thaliana chromosome 5. III. Sequence
RT features of the regions of 1,191,918 bp covered by seventeen physically
RT assigned P1 clones.";
RL DNA Res. 4:401-414(1997).
RN [3]
RP GENOME REANNOTATION.
RC STRAIN=cv. Columbia;
RX PubMed=27862469; DOI=10.1111/tpj.13415;
RA Cheng C.Y., Krishnakumar V., Chan A.P., Thibaud-Nissen F., Schobel S.,
RA Town C.D.;
RT "Araport11: a complete reannotation of the Arabidopsis thaliana reference
RT genome.";
RL Plant J. 89:789-804(2017).
RN [4]
RP NUCLEOTIDE SEQUENCE [LARGE SCALE MRNA].
RC STRAIN=cv. Columbia;
RX PubMed=14593172; DOI=10.1126/science.1088305;
RA Yamada K., Lim J., Dale J.M., Chen H., Shinn P., Palm C.J., Southwick A.M.,
RA Wu H.C., Kim C.J., Nguyen M., Pham P.K., Cheuk R.F., Karlin-Newmann G.,
RA Liu S.X., Lam B., Sakano H., Wu T., Yu G., Miranda M., Quach H.L.,
RA Tripp M., Chang C.H., Lee J.M., Toriumi M.J., Chan M.M., Tang C.C.,
RA Onodera C.S., Deng J.M., Akiyama K., Ansari Y., Arakawa T., Banh J.,
RA Banno F., Bowser L., Brooks S.Y., Carninci P., Chao Q., Choy N., Enju A.,
RA Goldsmith A.D., Gurjal M., Hansen N.F., Hayashizaki Y., Johnson-Hopson C.,
RA Hsuan V.W., Iida K., Karnes M., Khan S., Koesema E., Ishida J., Jiang P.X.,
RA Jones T., Kawai J., Kamiya A., Meyers C., Nakajima M., Narusaka M.,
RA Seki M., Sakurai T., Satou M., Tamse R., Vaysberg M., Wallender E.K.,
RA Wong C., Yamamura Y., Yuan S., Shinozaki K., Davis R.W., Theologis A.,
RA Ecker J.R.;
RT "Empirical analysis of transcriptional activity in the Arabidopsis
RT genome.";
RL Science 302:842-846(2003).
RN [5]
RP NUCLEOTIDE SEQUENCE [LARGE SCALE MRNA].
RA Brover V.V., Troukhan M.E., Alexandrov N.A., Lu Y.-P., Flavell R.B.,
RA Feldmann K.A.;
RT "Full-length cDNA from Arabidopsis thaliana.";
RL Submitted (MAR-2002) to the EMBL/GenBank/DDBJ databases.
RN [6]
RP FUNCTION, MUTAGENESIS OF CYS-197, ACTIVITY REGULATION, AND
RP BIOPHYSICOCHEMICAL PROPERTIES.
RX PubMed=11060302; DOI=10.1074/jbc.m007890200;
RA Piotrowski M., Schoenfelder S., Weiler E.W.;
RT "The Arabidopsis thaliana isogene NIT4 and its orthologs in tobacco encode
RT beta-cyano-L-alanine hydratase/nitrilase.";
RL J. Biol. Chem. 276:2616-2621(2001).
RN [7]
RP STRUCTURE BY ELECTRON MICROSCOPY (3.40 ANGSTROMS), AND SUBUNIT.
RX PubMed=31341959; DOI=10.1038/s42003-019-0505-4;
RA Mulelu A.E., Kirykowicz A.M., Woodward J.D.;
RT "Cryo-EM and directed evolution reveal how Arabidopsis nitrilase
RT specificity is influenced by its quaternary structure.";
RL Commun. Biol. 2:260-260(2019).
CC -!- FUNCTION: Highly specific for beta-cyano-L-alanine (Ala(CN)). Low
CC activity with 3-phenylpropionitrile (PPN) or allylcyanide and no
CC activity with indole-3-acetonitrile. Not associated with auxin
CC production but may be involved in cyanide detoxification.
CC {ECO:0000269|PubMed:11060302}.
CC -!- CATALYTIC ACTIVITY:
CC Reaction=a nitrile + 2 H2O = a carboxylate + NH4(+);
CC Xref=Rhea:RHEA:21724, ChEBI:CHEBI:15377, ChEBI:CHEBI:18379,
CC ChEBI:CHEBI:28938, ChEBI:CHEBI:29067; EC=3.5.5.1;
CC Evidence={ECO:0000255|PROSITE-ProRule:PRU10105};
CC -!- CATALYTIC ACTIVITY:
CC Reaction=3-cyano-L-alanine + 2 H2O = L-aspartate + NH4(+);
CC Xref=Rhea:RHEA:11188, ChEBI:CHEBI:15377, ChEBI:CHEBI:28938,
CC ChEBI:CHEBI:29991, ChEBI:CHEBI:77860; EC=3.5.5.4;
CC -!- CATALYTIC ACTIVITY:
CC Reaction=L-asparagine = 3-cyano-L-alanine + H2O; Xref=Rhea:RHEA:15385,
CC ChEBI:CHEBI:15377, ChEBI:CHEBI:58048, ChEBI:CHEBI:77860; EC=4.2.1.65;
CC -!- ACTIVITY REGULATION: Both catalytic activities are inhibited by N-
CC ethylmaleimide and are up-regulated during senescence.
CC {ECO:0000269|PubMed:11060302}.
CC -!- BIOPHYSICOCHEMICAL PROPERTIES:
CC Kinetic parameters:
CC KM=0.74 mM for Ala(CN) (for the nitrilase activity)
CC {ECO:0000269|PubMed:11060302};
CC KM=0.70 mM for Ala(CN) (for the nitrile hydratase activity)
CC {ECO:0000269|PubMed:11060302};
CC Vmax=1.84 umol/sec/mg enzyme for the nitrilase activity
CC {ECO:0000269|PubMed:11060302};
CC Vmax=2.55 umol/sec/mg enzyme for the nitrile hydratase activity
CC {ECO:0000269|PubMed:11060302};
CC pH dependence:
CC Optimum pH is 7-9 for both activities. {ECO:0000269|PubMed:11060302};
CC Temperature dependence:
CC Optimum temperature for both activities is 40 degrees Celsius.
CC {ECO:0000269|PubMed:11060302};
CC -!- SUBUNIT: Forms a homopentamer composed of five dimers.
CC {ECO:0000269|PubMed:31341959}.
CC -!- SUBCELLULAR LOCATION: Cell membrane; Peripheral membrane protein;
CC Cytoplasmic side. Note=Tightly associated with the plasma membrane.
CC -!- SIMILARITY: Belongs to the carbon-nitrogen hydrolase superfamily.
CC Nitrilase family. {ECO:0000305}.
CC -!- SEQUENCE CAUTION:
CC Sequence=AAM65906.1; Type=Erroneous initiation; Note=Truncated N-terminus.; Evidence={ECO:0000305};
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DR EMBL; U09961; AAA19628.1; -; mRNA.
DR EMBL; AB007651; BAB08328.1; -; Genomic_DNA.
DR EMBL; CP002688; AED93008.1; -; Genomic_DNA.
DR EMBL; AF372965; AAK50102.1; -; mRNA.
DR EMBL; AY124854; AAM70563.1; -; mRNA.
DR EMBL; AY088367; AAM65906.1; ALT_INIT; mRNA.
DR PIR; T52265; T52265.
DR RefSeq; NP_197622.1; NM_122135.6.
DR PDB; 6I00; EM; 3.40 A; A/B/C/D/E/F/G/H/I/J/K/L=1-355.
DR PDB; 6I5T; EM; 3.90 A; A/B/C/D/E/F=1-355.
DR PDB; 6I5U; EM; 3.90 A; A/B/C/D/E/F=1-355.
DR PDBsum; 6I00; -.
DR PDBsum; 6I5T; -.
DR PDBsum; 6I5U; -.
DR AlphaFoldDB; P46011; -.
DR SMR; P46011; -.
DR STRING; 3702.AT5G22300.1; -.
DR PaxDb; P46011; -.
DR PRIDE; P46011; -.
DR ProteomicsDB; 249446; -.
DR EnsemblPlants; AT5G22300.1; AT5G22300.1; AT5G22300.
DR GeneID; 832290; -.
DR Gramene; AT5G22300.1; AT5G22300.1; AT5G22300.
DR KEGG; ath:AT5G22300; -.
DR Araport; AT5G22300; -.
DR TAIR; locus:2176377; AT5G22300.
DR eggNOG; KOG0805; Eukaryota.
DR HOGENOM; CLU_030130_6_1_1; -.
DR InParanoid; P46011; -.
DR OMA; PKGLDFG; -.
DR OrthoDB; 1001274at2759; -.
DR PhylomeDB; P46011; -.
DR PRO; PR:P46011; -.
DR Proteomes; UP000006548; Chromosome 5.
DR ExpressionAtlas; P46011; baseline and differential.
DR Genevisible; P46011; AT.
DR GO; GO:0005829; C:cytosol; HDA:TAIR.
DR GO; GO:0005886; C:plasma membrane; IEA:UniProtKB-SubCell.
DR GO; GO:0047558; F:3-cyanoalanine hydratase activity; IDA:TAIR.
DR GO; GO:0047427; F:cyanoalanine nitrilase activity; IDA:TAIR.
DR GO; GO:0080061; F:indole-3-acetonitrile nitrilase activity; IDA:TAIR.
DR GO; GO:0000257; F:nitrilase activity; IDA:TAIR.
DR GO; GO:0018822; F:nitrile hydratase activity; IDA:TAIR.
DR GO; GO:0019499; P:cyanide metabolic process; IEP:TAIR.
DR GO; GO:0051410; P:detoxification of nitrogen compound; IEP:TAIR.
DR CDD; cd07564; nitrilases_CHs; 1.
DR Gene3D; 3.60.110.10; -; 1.
DR InterPro; IPR003010; C-N_Hydrolase.
DR InterPro; IPR036526; C-N_Hydrolase_sf.
DR InterPro; IPR000132; Nitrilase/CN_hydratase_CS.
DR InterPro; IPR044149; Nitrilases_CHs.
DR PANTHER; PTHR46044; PTHR46044; 1.
DR Pfam; PF00795; CN_hydrolase; 1.
DR SUPFAM; SSF56317; SSF56317; 1.
DR PROSITE; PS50263; CN_HYDROLASE; 1.
DR PROSITE; PS00920; NITRIL_CHT_1; 1.
DR PROSITE; PS00921; NITRIL_CHT_2; 1.
PE 1: Evidence at protein level;
KW 3D-structure; Acetylation; Cell membrane; Hydrolase; Lyase; Membrane;
KW Reference proteome.
FT INIT_MET 1
FT /note="Removed"
FT /evidence="ECO:0000250|UniProtKB:P32962"
FT CHAIN 2..355
FT /note="Bifunctional nitrilase/nitrile hydratase NIT4"
FT /id="PRO_0000204039"
FT DOMAIN 36..308
FT /note="CN hydrolase"
FT /evidence="ECO:0000255|PROSITE-ProRule:PRU00054"
FT ACT_SITE 76
FT /note="Proton acceptor"
FT /evidence="ECO:0000255|PROSITE-ProRule:PRU00054"
FT ACT_SITE 163
FT /note="Proton donor"
FT /evidence="ECO:0000255|PROSITE-ProRule:PRU00054"
FT ACT_SITE 197
FT /note="Nucleophile"
FT /evidence="ECO:0000255|PROSITE-ProRule:PRU00054,
FT ECO:0000255|PROSITE-ProRule:PRU10105"
FT MOD_RES 2
FT /note="N-acetylserine"
FT /evidence="ECO:0000250|UniProtKB:P32962"
FT MUTAGEN 197
FT /note="C->A: Loss of nitrilase activity and 95% reduction
FT of the nitrile hydratase activity."
FT /evidence="ECO:0000269|PubMed:11060302"
FT STRAND 36..40
FT /evidence="ECO:0007829|PDB:6I00"
FT TURN 50..52
FT /evidence="ECO:0007829|PDB:6I00"
FT HELIX 56..65
FT /evidence="ECO:0007829|PDB:6I00"
FT TURN 66..68
FT /evidence="ECO:0007829|PDB:6I00"
FT STRAND 71..73
FT /evidence="ECO:0007829|PDB:6I00"
FT TURN 76..80
FT /evidence="ECO:0007829|PDB:6I00"
FT STRAND 91..93
FT /evidence="ECO:0007829|PDB:6I00"
FT HELIX 97..107
FT /evidence="ECO:0007829|PDB:6I00"
FT STRAND 113..115
FT /evidence="ECO:0007829|PDB:6I00"
FT TURN 116..118
FT /evidence="ECO:0007829|PDB:6I00"
FT HELIX 119..126
FT /evidence="ECO:0007829|PDB:6I00"
FT TURN 127..129
FT /evidence="ECO:0007829|PDB:6I00"
FT STRAND 131..142
FT /evidence="ECO:0007829|PDB:6I00"
FT STRAND 144..151
FT /evidence="ECO:0007829|PDB:6I00"
FT STRAND 157..160
FT /evidence="ECO:0007829|PDB:6I00"
FT TURN 170..173
FT /evidence="ECO:0007829|PDB:6I00"
FT STRAND 190..194
FT /evidence="ECO:0007829|PDB:6I00"
FT STRAND 199..201
FT /evidence="ECO:0007829|PDB:6I00"
FT HELIX 203..209
FT /evidence="ECO:0007829|PDB:6I00"
FT HELIX 210..212
FT /evidence="ECO:0007829|PDB:6I00"
FT STRAND 215..220
FT /evidence="ECO:0007829|PDB:6I00"
FT HELIX 226..238
FT /evidence="ECO:0007829|PDB:6I00"
FT STRAND 241..246
FT /evidence="ECO:0007829|PDB:6I00"
FT STRAND 290..292
FT /evidence="ECO:0007829|PDB:6I00"
FT STRAND 302..306
FT /evidence="ECO:0007829|PDB:6I00"
FT TURN 312..317
FT /evidence="ECO:0007829|PDB:6I00"
FT TURN 320..323
FT /evidence="ECO:0007829|PDB:6I00"
FT TURN 327..329
FT /evidence="ECO:0007829|PDB:6I00"
FT STRAND 330..336
FT /evidence="ECO:0007829|PDB:6I00"
FT STRAND 341..346
FT /evidence="ECO:0007829|PDB:6I00"
SQ SEQUENCE 355 AA; 38895 MW; 8A042F7DFC5600F6 CRC64;
MSMQQETSHM TAAPQTNGHQ IFPEIDMSAG DSSSIVRATV VQASTVFYDT PATLDKAERL
LSEAAENGSQ LVVFPEAFIG GYPRGSTFEL AIGSRTAKGR DDFRKYHASA IDVPGPEVER
LALMAKKYKV YLVMGVIERE GYTLYCTVLF FDSQGLFLGK HRKLMPTALE RCIWGFGDGS
TIPVFDTPIG KIGAAICWEN RMPSLRTAMY AKGIEIYCAP TADSRETWLA SMTHIALEGG
CFVLSANQFC RRKDYPSPPE YMFSGSEESL TPDSVVCAGG SSIISPLGIV LAGPNYRGEA
LITADLDLGD IARAKFDFDV VGHYSRPEVF SLNIREHPRK AVSFKTSKVM EDESV
