ASNB_ECOLI
ID ASNB_ECOLI Reviewed; 554 AA.
AC P22106;
DT 01-AUG-1991, integrated into UniProtKB/Swiss-Prot.
DT 23-JAN-2007, sequence version 3.
DT 03-AUG-2022, entry version 178.
DE RecName: Full=Asparagine synthetase B [glutamine-hydrolyzing];
DE Short=AS-B;
DE EC=6.3.5.4;
GN Name=asnB; OrderedLocusNames=b0674, JW0660;
OS Escherichia coli (strain K12).
OC Bacteria; Proteobacteria; Gammaproteobacteria; Enterobacterales;
OC Enterobacteriaceae; Escherichia.
OX NCBI_TaxID=83333;
RN [1]
RP NUCLEOTIDE SEQUENCE [GENOMIC DNA].
RC STRAIN=K12;
RX PubMed=1973930; DOI=10.1016/s0021-9258(19)38244-4;
RA Scofield M.A., Lewis W.S., Schuster S.M.;
RT "Nucleotide sequence of Escherichia coli asnB and deduced amino acid
RT sequence of asparagine synthetase B.";
RL J. Biol. Chem. 265:12895-12902(1990).
RN [2]
RP NUCLEOTIDE SEQUENCE [LARGE SCALE GENOMIC DNA].
RC STRAIN=K12 / W3110 / ATCC 27325 / DSM 5911;
RX PubMed=8905232; DOI=10.1093/dnares/3.3.137;
RA Oshima T., Aiba H., Baba T., Fujita K., Hayashi K., Honjo A., Ikemoto K.,
RA Inada T., Itoh T., Kajihara M., Kanai K., Kashimoto K., Kimura S.,
RA Kitagawa M., Makino K., Masuda S., Miki T., Mizobuchi K., Mori H.,
RA Motomura K., Nakamura Y., Nashimoto H., Nishio Y., Saito N., Sampei G.,
RA Seki Y., Tagami H., Takemoto K., Wada C., Yamamoto Y., Yano M.,
RA Horiuchi T.;
RT "A 718-kb DNA sequence of the Escherichia coli K-12 genome corresponding to
RT the 12.7-28.0 min region on the linkage map.";
RL DNA Res. 3:137-155(1996).
RN [3]
RP NUCLEOTIDE SEQUENCE [LARGE SCALE GENOMIC DNA].
RC STRAIN=K12 / MG1655 / ATCC 47076;
RX PubMed=9278503; DOI=10.1126/science.277.5331.1453;
RA Blattner F.R., Plunkett G. III, Bloch C.A., Perna N.T., Burland V.,
RA Riley M., Collado-Vides J., Glasner J.D., Rode C.K., Mayhew G.F.,
RA Gregor J., Davis N.W., Kirkpatrick H.A., Goeden M.A., Rose D.J., Mau B.,
RA Shao Y.;
RT "The complete genome sequence of Escherichia coli K-12.";
RL Science 277:1453-1462(1997).
RN [4]
RP NUCLEOTIDE SEQUENCE [LARGE SCALE GENOMIC DNA].
RC STRAIN=K12 / W3110 / ATCC 27325 / DSM 5911;
RX PubMed=16738553; DOI=10.1038/msb4100049;
RA Hayashi K., Morooka N., Yamamoto Y., Fujita K., Isono K., Choi S.,
RA Ohtsubo E., Baba T., Wanner B.L., Mori H., Horiuchi T.;
RT "Highly accurate genome sequences of Escherichia coli K-12 strains MG1655
RT and W3110.";
RL Mol. Syst. Biol. 2:E1-E5(2006).
RN [5]
RP PROTEIN SEQUENCE OF N-TERMINUS, CLEAVAGE OF INITIATOR METHIONINE, FUNCTION,
RP CATALYTIC ACTIVITY, SUBSTRATE SPECIFICITY, BIOPHYSICOCHEMICAL PROPERTIES,
RP AND MUTAGENESIS OF CYS-2; HIS-30; ASP-34; HIS-81 AND ALA-105.
RC STRAIN=K12;
RX PubMed=7907328; DOI=10.1016/s0021-9258(17)37307-6;
RA Boehlein S.K., Richards N.G., Schuster S.M.;
RT "Glutamine-dependent nitrogen transfer in Escherichia coli asparagine
RT synthetase B. Searching for the catalytic triad.";
RL J. Biol. Chem. 269:7450-7457(1994).
RN [6]
RP FUNCTION IN ASPARAGINE BIOSYNTHESIS.
RC STRAIN=K12;
RX PubMed=6102982; DOI=10.1128/jb.142.1.212-220.1980;
RA Humbert R., Simoni R.D.;
RT "Genetic and biomedical studies demonstrating a second gene coding for
RT asparagine synthetase in Escherichia coli.";
RL J. Bacteriol. 142:212-220(1980).
RN [7]
RP ACTIVITY REGULATION, AND INHIBITION STUDIES.
RX PubMed=8691431; DOI=10.1021/jm9601009;
RA Parr I.B., Boehlein S.K., Dribben A.B., Schuster S.M., Richards N.G.;
RT "Mapping the aspartic acid binding site of Escherichia coli asparagine
RT synthetase B using substrate analogs.";
RL J. Med. Chem. 39:2367-2378(1996).
RN [8]
RP KINETIC PARAMETERS.
RC STRAIN=K12;
RX PubMed=12706338; DOI=10.1016/s0003-9861(03)00118-8;
RA Tesson A.R., Soper T.S., Ciustea M., Richards N.G.;
RT "Revisiting the steady state kinetic mechanism of glutamine-dependent
RT asparagine synthetase from Escherichia coli.";
RL Arch. Biochem. Biophys. 413:23-31(2003).
RN [9]
RP FUNCTION, CATALYTIC ACTIVITY, MUTAGENESIS OF GLU-349, AND KINETIC MODEL.
RC STRAIN=K12;
RX PubMed=20853825; DOI=10.1021/bi1010688;
RA Meyer M.E., Gutierrez J.A., Raushel F.M., Richards N.G.;
RT "A conserved glutamate controls the commitment to acyl-adenylate formation
RT in asparagine synthetase.";
RL Biochemistry 49:9391-9401(2010).
RN [10]
RP X-RAY CRYSTALLOGRAPHY (2.0 ANGSTROMS) OF 2-554 OF MUTANT ALA-2 IN COMPLEX
RP WITH AMP AND L-GLUTAMINE, AND SUBUNIT.
RC STRAIN=K12;
RX PubMed=10587437; DOI=10.1021/bi9915768;
RA Larsen T.M., Boehlein S.K., Schuster S.M., Richards N.G.J., Thoden J.B.,
RA Holden H.M., Rayment I.;
RT "Three-dimensional structure of Escherichia coli asparagine synthetase B: a
RT short journey from substrate to product.";
RL Biochemistry 38:16146-16157(1999).
RN [11]
RP ERRATUM OF PUBMED:10587437.
RX PubMed=10852734; DOI=10.1021/bi005109y;
RA Larsen T.M., Boehlein S.K., Schuster S.M., Richards N.G.J., Thoden J.B.,
RA Holden H.M., Rayment I.;
RL Biochemistry 39:7330-7330(2000).
CC -!- FUNCTION: Catalyzes the ATP-dependent conversion of aspartate into
CC asparagine, using glutamine as a source of nitrogen. Can also use
CC ammonia as the nitrogen source in vitro, albeit with lower efficiency.
CC As nucleotide substrates, ATP and dATP are utilized at a similar rate
CC in both the glutamine- and ammonia-dependent reactions, whereas GTP
CC utilization is only 15% that of ATP, and CTP, UTP, ITP and XTP are very
CC poor or not substrates. Also exhibits glutaminase activity.
CC {ECO:0000269|PubMed:20853825, ECO:0000269|PubMed:6102982,
CC ECO:0000269|PubMed:7907328}.
CC -!- CATALYTIC ACTIVITY:
CC Reaction=ATP + H2O + L-aspartate + L-glutamine = AMP + diphosphate +
CC H(+) + L-asparagine + L-glutamate; Xref=Rhea:RHEA:12228,
CC ChEBI:CHEBI:15377, ChEBI:CHEBI:15378, ChEBI:CHEBI:29985,
CC ChEBI:CHEBI:29991, ChEBI:CHEBI:30616, ChEBI:CHEBI:33019,
CC ChEBI:CHEBI:58048, ChEBI:CHEBI:58359, ChEBI:CHEBI:456215; EC=6.3.5.4;
CC Evidence={ECO:0000269|PubMed:20853825, ECO:0000269|PubMed:7907328};
CC -!- ACTIVITY REGULATION: Glutamine-dependent asparagine synthesis activity
CC can be inhibited by aspartic acid analogs (such as a sulfinate
CC derivative and (2S,3R)-2-amino-3-methylsuccinate) in vitro; the
CC inhibition is competitive with respect to aspartate.
CC {ECO:0000269|PubMed:8691431}.
CC -!- BIOPHYSICOCHEMICAL PROPERTIES:
CC Kinetic parameters:
CC KM=17 mM for ammonia {ECO:0000269|PubMed:12706338,
CC ECO:0000269|PubMed:7907328};
CC KM=0.53 mM for aspartate (when assaying the ammonia-dependent
CC synthetase reaction) {ECO:0000269|PubMed:12706338,
CC ECO:0000269|PubMed:7907328};
CC KM=0.85 mM for aspartate (when assaying the glutamine-dependent
CC synthetase reaction) {ECO:0000269|PubMed:12706338,
CC ECO:0000269|PubMed:7907328};
CC KM=0.26 mM for ATP (when assaying the glutamine-dependent synthetase
CC reaction) {ECO:0000269|PubMed:12706338, ECO:0000269|PubMed:7907328};
CC KM=0.66 mM for glutamine (when assaying the glutamine-dependent
CC synthetase reaction) {ECO:0000269|PubMed:12706338,
CC ECO:0000269|PubMed:7907328};
CC pH dependence:
CC Optimum pH is 6.5-8. {ECO:0000269|PubMed:7907328};
CC -!- PATHWAY: Amino-acid biosynthesis; L-asparagine biosynthesis; L-
CC asparagine from L-aspartate (L-Gln route): step 1/1.
CC -!- SUBUNIT: Homodimer. {ECO:0000269|PubMed:10587437}.
CC -!- INTERACTION:
CC P22106; P22106: asnB; NbExp=3; IntAct=EBI-549123, EBI-549123;
CC -!- SIMILARITY: Belongs to the asparagine synthetase family. {ECO:0000305}.
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DR EMBL; J05554; AAA23498.1; -; Genomic_DNA.
DR EMBL; U00096; AAC73768.1; -; Genomic_DNA.
DR EMBL; AP009048; BAA35317.1; -; Genomic_DNA.
DR PIR; A36616; AJECN.
DR RefSeq; NP_415200.1; NC_000913.3.
DR RefSeq; WP_000337077.1; NZ_SSZK01000045.1.
DR PDB; 1CT9; X-ray; 2.00 A; A/B/C/D=2-554.
DR PDBsum; 1CT9; -.
DR AlphaFoldDB; P22106; -.
DR SMR; P22106; -.
DR BioGRID; 4261812; 416.
DR BioGRID; 849659; 1.
DR DIP; DIP-9177N; -.
DR IntAct; P22106; 7.
DR STRING; 511145.b0674; -.
DR MEROPS; C44.976; -.
DR jPOST; P22106; -.
DR PaxDb; P22106; -.
DR PRIDE; P22106; -.
DR EnsemblBacteria; AAC73768; AAC73768; b0674.
DR EnsemblBacteria; BAA35317; BAA35317; BAA35317.
DR GeneID; 60901229; -.
DR GeneID; 945281; -.
DR KEGG; ecj:JW0660; -.
DR KEGG; eco:b0674; -.
DR PATRIC; fig|1411691.4.peg.1605; -.
DR EchoBASE; EB0090; -.
DR eggNOG; COG0367; Bacteria.
DR HOGENOM; CLU_014658_2_2_6; -.
DR InParanoid; P22106; -.
DR OMA; DWSGIYS; -.
DR PhylomeDB; P22106; -.
DR BioCyc; EcoCyc:ASNSYNB-MON; -.
DR BioCyc; MetaCyc:ASNSYNB-MON; -.
DR BRENDA; 6.3.5.4; 2026.
DR UniPathway; UPA00134; UER00195.
DR EvolutionaryTrace; P22106; -.
DR PRO; PR:P22106; -.
DR Proteomes; UP000000318; Chromosome.
DR Proteomes; UP000000625; Chromosome.
DR GO; GO:0005737; C:cytoplasm; IDA:EcoliWiki.
DR GO; GO:0005829; C:cytosol; IDA:EcoCyc.
DR GO; GO:0016597; F:amino acid binding; IDA:UniProtKB.
DR GO; GO:0004066; F:asparagine synthase (glutamine-hydrolyzing) activity; IDA:UniProtKB.
DR GO; GO:0004071; F:aspartate-ammonia ligase activity; IDA:UniProtKB.
DR GO; GO:0005524; F:ATP binding; IDA:UniProtKB.
DR GO; GO:0042802; F:identical protein binding; IPI:IntAct.
DR GO; GO:0006529; P:asparagine biosynthetic process; IDA:UniProtKB.
DR GO; GO:0008652; P:cellular amino acid biosynthetic process; IMP:EcoliWiki.
DR GO; GO:0009063; P:cellular amino acid catabolic process; IMP:EcoliWiki.
DR GO; GO:0006541; P:glutamine metabolic process; IMP:EcoliWiki.
DR GO; GO:0070981; P:L-asparagine biosynthetic process; IEA:UniProtKB-UniPathway.
DR CDD; cd01991; Asn_Synthase_B_C; 1.
DR CDD; cd00712; AsnB; 1.
DR Gene3D; 3.40.50.620; -; 1.
DR Gene3D; 3.60.20.10; -; 1.
DR InterPro; IPR006426; Asn_synth_AEB.
DR InterPro; IPR001962; Asn_synthase.
DR InterPro; IPR033738; AsnB_N.
DR InterPro; IPR017932; GATase_2_dom.
DR InterPro; IPR029055; Ntn_hydrolases_N.
DR InterPro; IPR014729; Rossmann-like_a/b/a_fold.
DR Pfam; PF00733; Asn_synthase; 1.
DR Pfam; PF13537; GATase_7; 1.
DR PIRSF; PIRSF001589; Asn_synthetase_glu-h; 1.
DR SUPFAM; SSF56235; SSF56235; 1.
DR TIGRFAMs; TIGR01536; asn_synth_AEB; 1.
DR PROSITE; PS51278; GATASE_TYPE_2; 1.
PE 1: Evidence at protein level;
KW 3D-structure; Amino-acid biosynthesis; Asparagine biosynthesis;
KW ATP-binding; Direct protein sequencing; Glutamine amidotransferase; Ligase;
KW Nucleotide-binding; Reference proteome.
FT INIT_MET 1
FT /note="Removed"
FT /evidence="ECO:0000269|PubMed:7907328"
FT CHAIN 2..554
FT /note="Asparagine synthetase B [glutamine-hydrolyzing]"
FT /id="PRO_0000056931"
FT DOMAIN 2..186
FT /note="Glutamine amidotransferase type-2"
FT /evidence="ECO:0000255|PROSITE-ProRule:PRU00609"
FT ACT_SITE 2
FT /note="For GATase activity"
FT BINDING 50..54
FT /ligand="L-glutamine"
FT /ligand_id="ChEBI:CHEBI:58359"
FT BINDING 75..77
FT /ligand="L-glutamine"
FT /ligand_id="ChEBI:CHEBI:58359"
FT BINDING 99
FT /ligand="L-glutamine"
FT /ligand_id="ChEBI:CHEBI:58359"
FT BINDING 233
FT /ligand="ATP"
FT /ligand_id="ChEBI:CHEBI:30616"
FT BINDING 273
FT /ligand="ATP"
FT /ligand_id="ChEBI:CHEBI:30616"
FT BINDING 347..348
FT /ligand="ATP"
FT /ligand_id="ChEBI:CHEBI:30616"
FT SITE 349
FT /note="Important for beta-aspartyl-AMP intermediate
FT formation"
FT MUTAGEN 2
FT /note="C->A,S: Loss of glutamine-dependent activity but no
FT effect on ammonia-dependent asparagine synthetase
FT activity."
FT /evidence="ECO:0000269|PubMed:7907328"
FT MUTAGEN 30
FT /note="H->A: 4,5-fold decrease in glutamine affinity."
FT /evidence="ECO:0000269|PubMed:7907328"
FT MUTAGEN 34
FT /note="D->N,E: Little effect on the kinetic properties."
FT /evidence="ECO:0000269|PubMed:7907328"
FT MUTAGEN 81
FT /note="H->A: 5-fold decrease in glutamine affinity."
FT /evidence="ECO:0000269|PubMed:7907328"
FT MUTAGEN 105
FT /note="A->H: Little effect on the kinetic properties."
FT /evidence="ECO:0000269|PubMed:7907328"
FT MUTAGEN 349
FT /note="E->A,Q: Loss of glutamine- and ammonia-dependent
FT synthetase activity, but still exhibits glutaminase
FT activity."
FT /evidence="ECO:0000269|PubMed:20853825"
FT MUTAGEN 349
FT /note="E->D: 5-fold increase in affinity for aspartate when
FT assaying both the glutamine- and ammonia-dependent
FT synthetase reactions, and 2-fold decrease in kcat for these
FT reactions. Modifies the product glutamate/asparagine
FT stoichiometry."
FT /evidence="ECO:0000269|PubMed:20853825"
FT STRAND 3..9
FT /evidence="ECO:0007829|PDB:1CT9"
FT HELIX 14..26
FT /evidence="ECO:0007829|PDB:1CT9"
FT HELIX 27..31
FT /evidence="ECO:0007829|PDB:1CT9"
FT STRAND 35..40
FT /evidence="ECO:0007829|PDB:1CT9"
FT STRAND 42..50
FT /evidence="ECO:0007829|PDB:1CT9"
FT TURN 56..58
FT /evidence="ECO:0007829|PDB:1CT9"
FT STRAND 61..64
FT /evidence="ECO:0007829|PDB:1CT9"
FT STRAND 70..78
FT /evidence="ECO:0007829|PDB:1CT9"
FT HELIX 81..88
FT /evidence="ECO:0007829|PDB:1CT9"
FT TURN 89..91
FT /evidence="ECO:0007829|PDB:1CT9"
FT HELIX 99..101
FT /evidence="ECO:0007829|PDB:1CT9"
FT HELIX 102..110
FT /evidence="ECO:0007829|PDB:1CT9"
FT TURN 111..114
FT /evidence="ECO:0007829|PDB:1CT9"
FT HELIX 115..117
FT /evidence="ECO:0007829|PDB:1CT9"
FT STRAND 120..128
FT /evidence="ECO:0007829|PDB:1CT9"
FT TURN 129..132
FT /evidence="ECO:0007829|PDB:1CT9"
FT STRAND 133..138
FT /evidence="ECO:0007829|PDB:1CT9"
FT STRAND 147..150
FT /evidence="ECO:0007829|PDB:1CT9"
FT STRAND 156..161
FT /evidence="ECO:0007829|PDB:1CT9"
FT HELIX 162..164
FT /evidence="ECO:0007829|PDB:1CT9"
FT TURN 165..168
FT /evidence="ECO:0007829|PDB:1CT9"
FT STRAND 170..174
FT /evidence="ECO:0007829|PDB:1CT9"
FT STRAND 179..182
FT /evidence="ECO:0007829|PDB:1CT9"
FT TURN 183..185
FT /evidence="ECO:0007829|PDB:1CT9"
FT STRAND 187..190
FT /evidence="ECO:0007829|PDB:1CT9"
FT HELIX 195..197
FT /evidence="ECO:0007829|PDB:1CT9"
FT HELIX 199..202
FT /evidence="ECO:0007829|PDB:1CT9"
FT HELIX 209..223
FT /evidence="ECO:0007829|PDB:1CT9"
FT STRAND 230..233
FT /evidence="ECO:0007829|PDB:1CT9"
FT HELIX 238..250
FT /evidence="ECO:0007829|PDB:1CT9"
FT STRAND 269..275
FT /evidence="ECO:0007829|PDB:1CT9"
FT HELIX 279..291
FT /evidence="ECO:0007829|PDB:1CT9"
FT STRAND 294..299
FT /evidence="ECO:0007829|PDB:1CT9"
FT HELIX 302..316
FT /evidence="ECO:0007829|PDB:1CT9"
FT HELIX 321..339
FT /evidence="ECO:0007829|PDB:1CT9"
FT STRAND 344..346
FT /evidence="ECO:0007829|PDB:1CT9"
FT HELIX 351..355
FT /evidence="ECO:0007829|PDB:1CT9"
FT HELIX 359..363
FT /evidence="ECO:0007829|PDB:1CT9"
FT HELIX 367..380
FT /evidence="ECO:0007829|PDB:1CT9"
FT HELIX 381..383
FT /evidence="ECO:0007829|PDB:1CT9"
FT HELIX 385..394
FT /evidence="ECO:0007829|PDB:1CT9"
FT TURN 395..397
FT /evidence="ECO:0007829|PDB:1CT9"
FT STRAND 399..401
FT /evidence="ECO:0007829|PDB:1CT9"
FT HELIX 403..405
FT /evidence="ECO:0007829|PDB:1CT9"
FT HELIX 407..415
FT /evidence="ECO:0007829|PDB:1CT9"
FT HELIX 418..420
FT /evidence="ECO:0007829|PDB:1CT9"
FT HELIX 431..437
FT /evidence="ECO:0007829|PDB:1CT9"
FT HELIX 438..440
FT /evidence="ECO:0007829|PDB:1CT9"
FT HELIX 443..446
FT /evidence="ECO:0007829|PDB:1CT9"
FT HELIX 460..472
FT /evidence="ECO:0007829|PDB:1CT9"
FT HELIX 475..479
FT /evidence="ECO:0007829|PDB:1CT9"
FT HELIX 481..484
FT /evidence="ECO:0007829|PDB:1CT9"
FT HELIX 493..505
FT /evidence="ECO:0007829|PDB:1CT9"
FT HELIX 509..514
FT /evidence="ECO:0007829|PDB:1CT9"
SQ SEQUENCE 554 AA; 62659 MW; 9091B269112C9F5C CRC64;
MCSIFGVFDI KTDAVELRKK ALELSRLMRH RGPDWSGIYA SDNAILAHER LSIVDVNAGA
QPLYNQQKTH VLAVNGEIYN HQALRAEYGD RYQFQTGSDC EVILALYQEK GPEFLDDLQG
MFAFALYDSE KDAYLIGRDH LGIIPLYMGY DEHGQLYVAS EMKALVPVCR TIKEFPAGSY
LWSQDGEIRS YYHRDWFDYD AVKDNVTDKN ELRQALEDSV KSHLMSDVPY GVLLSGGLDS
SIISAITKKY AARRVEDQER SEAWWPQLHS FAVGLPGSPD LKAAQEVANH LGTVHHEIHF
TVQEGLDAIR DVIYHIETYD VTTIRASTPM YLMSRKIKAM GIKMVLSGEG SDEVFGGYLY
FHKAPNAKEL HEETVRKLLA LHMYDCARAN KAMSAWGVEA RVPFLDKKFL DVAMRINPQD
KMCGNGKMEK HILRECFEAY LPASVAWRQK EQFSDGVGYS WIDTLKEVAA QQVSDQQLET
ARFRFPYNTP TSKEAYLYRE IFEELFPLPS AAECVPGGPS VACSSAKAIE WDEAFKKMDD
PSGRAVGVHQ SAYK
