P22106 (ASNB_ECOLI) Reviewed, UniProtKB/Swiss-Prot

Names and origin

Protein names

Recommended name:
Asparagine synthetase B [glutamine-hydrolyzing]
Short name=AS-B
EC=6.3.5.4

Gene names

Name:	asnB
Ordered Locus Names:	b0674, JW0660

Organism

Escherichia coli (strain K12) [Reference proteome] [HAMAP]

Taxonomic identifier

83333 [NCBI]

Taxonomic lineage

cellular organisms › Bacteria › Proteobacteria › Gammaproteobacteria › Enterobacteriales › Enterobacteriaceae › Escherichia › Escherichia coli

Protein attributes

Sequence length	554 AA.
Sequence status	Complete.
Sequence processing	The displayed sequence is further processed into a mature form.
Protein existence	Evidence at protein level

General annotation (Comments)

Function	Catalyzes the ATP-dependent conversion of aspartate into asparagine, using glutamine as a source of nitrogen. Can also use ammonia as the nitrogen source in vitro, albeit with lower efficiency. As nucleotide substrates, ATP and dATP are utilized at a similar rate in both the glutamine- and ammonia-dependent reactions, whereas GTP utilization is only 15% that of ATP, and CTP, UTP, ITP and XTP are very poor or not substrates. Also exhibits glutaminase activity. Ref.5 Ref.6 Ref.9
Catalytic activity	ATP + L-aspartate + L-glutamine + H2O = AMP + diphosphate + L-asparagine + L-glutamate. Ref.5 Ref.9
Enzyme regulation	Glutamine-dependent asparagine synthesis activity can be inhibited by aspartic acid analogs (such as a sulfinate derivative and (2S,3R)-2-amino-3-methylsuccinate) in vitro; the inhibition is competitive with respect to aspartate. Ref.7
Pathway	Amino-acid biosynthesis; L-asparagine biosynthesis; L-asparagine from L-aspartate (L-Gln route): step 1/1.
Subunit structure	Homodimer. Ref.10
Sequence similarities	Belongs to the asparagine synthetase family. Contains 1 glutamine amidotransferase type-2 domain.
Biophysicochemical properties	Kinetic parameters: KM=17 mM for ammonia Ref.5 Ref.8 Ref.9 KM=0.53 mM for aspartate (when assaying the ammonia-dependent synthetase reaction) KM=0.85 mM for aspartate (when assaying the glutamine-dependent synthetase reaction) KM=0.26 mM for ATP (when assaying the glutamine-dependent synthetase reaction) KM=0.66 mM for glutamine (when assaying the glutamine-dependent synthetase reaction) pH dependence: Optimum pH is 6.5-8.

Ontologies

Keywords
Biological process	Amino-acid biosynthesis Asparagine biosynthesis
Domain	Glutamine amidotransferase
Ligand	ATP-binding Nucleotide-binding
Molecular function	Ligase
Technical term	3D-structure Complete proteome Direct protein sequencing Reference proteome
Gene Ontology (GO)
Biological_process	L-asparagine biosynthetic process Inferred from electronic annotation. Source: UniProtKB-UniPathway asparagine biosynthetic process Inferred from direct assay Ref.5. Source: UniProtKB cellular amino acid catabolic process Inferred from mutant phenotype Ref.6. Source: EcoliWiki glutamine metabolic process Inferred from mutant phenotype Ref.6. Source: EcoliWiki
Cellular_component	cytoplasm Inferred from direct assay PubMed 4895361Ref.6. Source: EcoliWiki
Molecular_function	ATP binding Inferred from direct assay Ref.5. Source: UniProtKB amino acid binding Inferred from direct assay Ref.5. Source: UniProtKB asparagine synthase (glutamine-hydrolyzing) activity Inferred from direct assay Ref.5. Source: UniProtKB aspartate-ammonia ligase activity Inferred from direct assay Ref.5. Source: UniProtKB
Complete GO annotation...

Sequence annotation (Features)

Feature key

Position(s)

Length

Description

Graphical view

Feature identifier

Molecule processing

Initiator methionine

1

1

Removed Ref.5

Chain

2 – 554

553

Asparagine synthetase B [glutamine-hydrolyzing]

PRO_0000056931

Regions

Domain

2 – 186

185

Glutamine amidotransferase type-2

Nucleotide binding

347 – 348

2

ATP

Region

50 – 54

5

Glutamine binding

Region

75 – 77

3

Glutamine binding

Sites

Active site

2

1

For GATase activity

Binding site

99

1

Glutamine

Binding site

233

1

ATP; via carbonyl oxygen

Binding site

273

1

ATP; via amide nitrogen and carbonyl oxygen

Site

349

1

Important for beta-aspartyl-AMP intermediate formation

Experimental info

Mutagenesis

2

1

C → A or S: Loss of glutamine-dependent activity but no effect on ammonia-dependent asparagine synthetase activity. Ref.5

Mutagenesis

30

1

H → A: 4,5-fold decrease in glutamine affinity. Ref.5

Mutagenesis

34

1

D → N or E: Little effect on the kinetic properties. Ref.5

Mutagenesis

81

1

H → A: 5-fold decrease in glutamine affinity. Ref.5

Mutagenesis

105

1

A → H: Little effect on the kinetic properties. Ref.5

Mutagenesis

349

1

E → A or Q: Loss of glutamine- and ammonia-dependent synthetase activity, but still exhibits glutaminase activity. Ref.9

Mutagenesis

349

1

E → D: 5-fold increase in affinity for asparate when assaying both the glutamine- and ammonia-dependent synthetase reactions, and 2-fold decrease in kcat for these reactions. Modifies the product glutamate/asparagine stoichiometry. Ref.9

Secondary structure

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554

Helix Strand Turn

Details...

Beta strand

3 – 9

7

Helix

14 – 26

13

Helix

27 – 31

5

Beta strand

35 – 40

6

Beta strand

42 – 50

9

Turn

56 – 58

3

Beta strand

61 – 64

4

Beta strand

70 – 78

9

Helix

81 – 88

8

Turn

89 – 91

3

Helix

99 – 101

3

Helix

102 – 110

9

Turn

111 – 114

4

Helix

115 – 117

3

Beta strand

120 – 128

9

Turn

129 – 132

4

Beta strand

133 – 138

6

Beta strand

147 – 150

4

Beta strand

156 – 161

6

Helix

162 – 164

3

Turn

165 – 168

4

Beta strand

170 – 174

5

Beta strand

179 – 182

4

Turn

183 – 185

3

Beta strand

187 – 190

4

Helix

195 – 197

3

Helix

199 – 202

4

Helix

209 – 223

15

Beta strand

230 – 233

4

Helix

238 – 250

13

Beta strand

269 – 275

7

Helix

279 – 291

13

Beta strand

294 – 299

6

Helix

302 – 316

15

Helix

321 – 339

19

Beta strand

344 – 346

3

Helix

351 – 355

5

Helix

359 – 363

5

Helix

367 – 380

14

Helix

381 – 383

3

Helix

385 – 394

10

Turn

395 – 397

3

Beta strand

399 – 401

3

Helix

403 – 405

3

Helix

407 – 415

9

Helix

418 – 420

3

Helix

431 – 437

7

Helix

438 – 440

3

Helix

443 – 446

4

Helix

460 – 472

13

Helix

475 – 479

5

Helix

481 – 484

4

Helix

493 – 505

13

Helix

509 – 514

6

Sequences

	Sequence		Length	Mass (Da)
	P22106 [UniParc]. Last modified January 23, 2007. Version 3. Checksum: 9091B269112C9F5C	FASTA	554	62,659
	10 20 30 40 50 60 MCSIFGVFDI KTDAVELRKK ALELSRLMRH RGPDWSGIYA SDNAILAHER LSIVDVNAGA 70 80 90 100 110 120 QPLYNQQKTH VLAVNGEIYN HQALRAEYGD RYQFQTGSDC EVILALYQEK GPEFLDDLQG 130 140 150 160 170 180 MFAFALYDSE KDAYLIGRDH LGIIPLYMGY DEHGQLYVAS EMKALVPVCR TIKEFPAGSY 190 200 210 220 230 240 LWSQDGEIRS YYHRDWFDYD AVKDNVTDKN ELRQALEDSV KSHLMSDVPY GVLLSGGLDS 250 260 270 280 290 300 SIISAITKKY AARRVEDQER SEAWWPQLHS FAVGLPGSPD LKAAQEVANH LGTVHHEIHF 310 320 330 340 350 360 TVQEGLDAIR DVIYHIETYD VTTIRASTPM YLMSRKIKAM GIKMVLSGEG SDEVFGGYLY 370 380 390 400 410 420 FHKAPNAKEL HEETVRKLLA LHMYDCARAN KAMSAWGVEA RVPFLDKKFL DVAMRINPQD 430 440 450 460 470 480 KMCGNGKMEK HILRECFEAY LPASVAWRQK EQFSDGVGYS WIDTLKEVAA QQVSDQQLET 490 500 510 520 530 540 ARFRFPYNTP TSKEAYLYRE IFEELFPLPS AAECVPGGPS VACSSAKAIE WDEAFKKMDD 550 PSGRAVGVHQ SAYK

References

	« Hide 'large scale' referencesShow 'large scale' references »
[1]	"Nucleotide sequence of Escherichia coli asnB and deduced amino acid sequence of asparagine synthetase B." Scofield M.A., Lewis W.S., Schuster S.M. J. Biol. Chem. 265:12895-12902(1990) [PubMed] [Europe PMC] [Abstract] Cited for: NUCLEOTIDE SEQUENCE [GENOMIC DNA]. Strain: K12.
[2]	"A 718-kb DNA sequence of the Escherichia coli K-12 genome corresponding to the 12.7-28.0 min region on the linkage map." Oshima T., Aiba H., Baba T., Fujita K., Hayashi K., Honjo A., Ikemoto K., Inada T., Itoh T., Kajihara M., Kanai K., Kashimoto K., Kimura S., Kitagawa M., Makino K., Masuda S., Miki T., Mizobuchi K. , Mori H., Motomura K., Nakamura Y., Nashimoto H., Nishio Y., Saito N., Sampei G., Seki Y., Tagami H., Takemoto K., Wada C., Yamamoto Y., Yano M., Horiuchi T. DNA Res. 3:137-155(1996) [PubMed] [Europe PMC] [Abstract] Cited for: NUCLEOTIDE SEQUENCE [LARGE SCALE GENOMIC DNA]. Strain: K12 / W3110 / ATCC 27325 / DSM 5911.
[3]	"The complete genome sequence of Escherichia coli K-12." Blattner F.R., Plunkett G. III, Bloch C.A., Perna N.T., Burland V., Riley M., Collado-Vides J., Glasner J.D., Rode C.K., Mayhew G.F., Gregor J., Davis N.W., Kirkpatrick H.A., Goeden M.A., Rose D.J., Mau B., Shao Y. Science 277:1453-1474(1997) [PubMed] [Europe PMC] [Abstract] Cited for: NUCLEOTIDE SEQUENCE [LARGE SCALE GENOMIC DNA]. Strain: K12 / MG1655 / ATCC 47076.
[4]	"Highly accurate genome sequences of Escherichia coli K-12 strains MG1655 and W3110." Hayashi K., Morooka N., Yamamoto Y., Fujita K., Isono K., Choi S., Ohtsubo E., Baba T., Wanner B.L., Mori H., Horiuchi T. Mol. Syst. Biol. 2:E1-E5(2006) [PubMed] [Europe PMC] [Abstract] Cited for: NUCLEOTIDE SEQUENCE [LARGE SCALE GENOMIC DNA]. Strain: K12 / W3110 / ATCC 27325 / DSM 5911.
[5]	"Glutamine-dependent nitrogen transfer in Escherichia coli asparagine synthetase B. Searching for the catalytic triad." Boehlein S.K., Richards N.G., Schuster S.M. J. Biol. Chem. 269:7450-7457(1994) [PubMed] [Europe PMC] [Abstract] Cited for: PROTEIN SEQUENCE OF N-TERMINUS, CLEAVAGE OF INITIATOR METHIONINE, FUNCTION, CATALYTIC ACTIVITY, SUBSTRATE SPECIFICITY, BIOPHYSICOCHEMICAL PROPERTIES, MUTAGENESIS OF CYS-2; HIS-30; ASP-34; HIS-81 AND ALA-105. Strain: K12.
[6]	"Genetic and biomedical studies demonstrating a second gene coding for asparagine synthetase in Escherichia coli." Humbert R., Simoni R.D. J. Bacteriol. 142:212-220(1980) [PubMed] [Europe PMC] [Abstract] Cited for: FUNCTION IN ASPARAGINE BIOSYNTHESIS. Strain: K12.
[7]	"Mapping the aspartic acid binding site of Escherichia coli asparagine synthetase B using substrate analogs." Parr I.B., Boehlein S.K., Dribben A.B., Schuster S.M., Richards N.G. J. Med. Chem. 39:2367-2378(1996) [PubMed] [Europe PMC] [Abstract] Cited for: ENZYME REGULATION, INHIBITION STUDIES.
[8]	"Revisiting the steady state kinetic mechanism of glutamine-dependent asparagine synthetase from Escherichia coli." Tesson A.R., Soper T.S., Ciustea M., Richards N.G. Arch. Biochem. Biophys. 413:23-31(2003) [PubMed] [Europe PMC] [Abstract] Cited for: KINETIC MECHANISM. Strain: K12.
[9]	"A conserved glutamate controls the commitment to acyl-adenylate formation in asparagine synthetase." Meyer M.E., Gutierrez J.A., Raushel F.M., Richards N.G. Biochemistry 49:9391-9401(2010) [PubMed] [Europe PMC] [Abstract] Cited for: FUNCTION, CATALYTIC ACTIVITY, MUTAGENESIS OF GLU-349, KINETIC MODEL. Strain: K12.
[10]	"Three-dimensional structure of Escherichia coli asparagine synthetase B: a short journey from substrate to product." Larsen T.M., Boehlein S.K., Schuster S.M., Richards N.G.J., Thoden J.B., Holden H.M., Rayment I. Biochemistry 38:16146-16157(1999) [PubMed] [Europe PMC] [Abstract] Cited for: X-RAY CRYSTALLOGRAPHY (2.0 ANGSTROMS) OF 2-554 OF MUTANT ALA-2 IN COMPLEX WITH AMP AND L-GLUTAMINE, SUBUNIT. Strain: K12.
[11]	Erratum Larsen T.M., Boehlein S.K., Schuster S.M., Richards N.G.J., Thoden J.B., Holden H.M., Rayment I. Biochemistry 39:7330-7330(2000) [PubMed] [Europe PMC] [Abstract]
+	Additional computationally mapped references.

Cross-references

Sequence databases

EMBL
GenBank
DDBJ

J05554 Genomic DNA. Translation: AAA23498.1.
U00096 Genomic DNA. Translation: AAC73768.1.
AP009048 Genomic DNA. Translation: BAA35317.1.

PIR

AJECN. A36616.

RefSeq

NP_415200.1. NC_000913.2.
YP_488954.1. NC_007779.1.

3D structure databases

PDBe
RCSB PDB
PDBj

Entry	Method	Resolution (Å)	Chain	Positions	PDBsum
1CT9	X-ray	2.00	A/B/C/D	3-554	[»]

ProteinModelPortal

P22106.

SMR

P22106. Positions 3-517.

ModBase

Search...

Protein-protein interaction databases

DIP

DIP-9177N.

IntAct

P22106. 7 interactions.

MINT

MINT-1306603.

STRING

511145.b0674.

Protein family/group databases

MEROPS

C44.976.

Proteomic databases

PaxDb

P22106.

PRIDE

P22106.

Protocols and materials databases

StructuralBiologyKnowledgebase

Search...

Genome annotation databases

EnsemblBacteria

AAC73768; AAC73768; b0674.
BAA35317; BAA35317; BAA35317.

GeneID

12932818.
945281.

KEGG

ecj:Y75_p0653.
eco:b0674.

PATRIC

32116529. VBIEscCol129921_0697.

Organism-specific databases

EchoBASE

EB0090.

EcoGene

EG10092. asnB.

Phylogenomic databases

eggNOG

COG0367.

HOGENOM

HOG000027493.

KO

K01953.

OMA

DEHGNYY.

ProtClustDB

PRK09431.

Enzyme and pathway databases

BioCyc

EcoCyc:ASNSYNB-MONOMER.
ECOL316407:JW0660-MONOMER.
MetaCyc:ASNSYNB-MONOMER.

UniPathway

UPA00134; UER00195.

Gene expression databases

Genevestigator

P22106.

Family and domain databases

Gene3D

3.40.50.620. 1 hit.

InterPro

IPR006426. Asn_synth_AEB.
IPR001962. Asn_synthase.
IPR017932. GATase_2_dom.
IPR000583. GATase_dom.
IPR014729. Rossmann-like_a/b/a_fold.
[Graphical view]

Pfam

PF00733. Asn_synthase. 1 hit.
PF13537. GATase_7. 1 hit.
[Graphical view]

PIRSF

PIRSF001589. Asn_synthetase_glu-h. 1 hit.

TIGRFAMs

TIGR01536. asn_synth_AEB. 1 hit.

PROSITE

PS51278. GATASE_TYPE_2. 1 hit.
[Graphical view]

ProtoNet

Search...

Other

DrugBank

DB00131. Adenosine monophosphate.

EvolutionaryTrace

P22106.

Entry information

Entry name

ASNB_ECOLI

Accession

Primary (citable) accession number: P22106

Entry history

Integrated into UniProtKB/Swiss-Prot:	August 1, 1991
Last sequence update:	January 23, 2007
Last modified:	May 1, 2013
This is version 120 of the entry and version 3 of the sequence. [Complete history]

Entry status

Reviewed (UniProtKB/Swiss-Prot)

Annotation program

Prokaryotic Protein Annotation Program