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 27-MAR-2024, entry version 186. 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; Pseudomonadota; 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}. CC --------------------------------------------------------------------------- CC Copyrighted by the UniProt Consortium, see https://www.uniprot.org/terms CC Distributed under the Creative Commons Attribution (CC BY 4.0) License CC --------------------------------------------------------------------------- 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; 511145-b0674; -. DR EnsemblBacteria; AAC73768; AAC73768; b0674. 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; GIVCAFD; -. DR OrthoDB; 9763290at2; -. DR PhylomeDB; P22106; -. DR BioCyc; EcoCyc:ASNSYNB-MONOMER; -. DR BioCyc; MetaCyc:ASNSYNB-MONOMER; -. 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:0042803; F:protein homodimerization activity; IPI:EcoCyc. DR GO; GO:0008652; P:amino acid biosynthetic process; IMP:EcoliWiki. DR GO; GO:0009063; P:amino acid catabolic process; IMP:EcoliWiki. DR GO; GO:0006529; P:asparagine biosynthetic process; IDA:UniProtKB. 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; 3.60.20.10; Glutamine Phosphoribosylpyrophosphate, subunit 1, domain 1; 1. DR Gene3D; 3.40.50.620; HUPs; 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 NCBIfam; TIGR01536; asn_synth_AEB; 1. DR PANTHER; PTHR11772; ASPARAGINE SYNTHETASE; 1. DR PANTHER; PTHR11772:SF2; ASPARAGINE SYNTHETASE [GLUTAMINE-HYDROLYZING]; 1. DR Pfam; PF00733; Asn_synthase; 1. DR Pfam; PF13537; GATase_7; 1. DR PIRSF; PIRSF001589; Asn_synthetase_glu-h; 1. DR SUPFAM; SSF52402; Adenine nucleotide alpha hydrolases-like; 1. DR SUPFAM; SSF56235; N-terminal nucleophile aminohydrolases (Ntn hydrolases); 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 //