EzCatDB: S00466
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DB codeS00466
RLCP classification3.103.70020.355 : Transfer
CATH domainDomain 13.90.550.10 : Spore Coat Polysaccharide Biosynthesis Protein SpsA; Chain ACatalytic domain
E.C.2.7.7.43

CATH domainRelated DB codes (homologues)
3.90.550.10 : Spore Coat Polysaccharide Biosynthesis Protein SpsA; Chain AS00709,S00465,D00417,D00859,D00860,T00415

Enzyme Name
UniProtKBKEGG

P0A0Z8
Protein nameN-acylneuraminate cytidylyltransferaseN-acylneuraminate cytidylyltransferase
CMP-sialate pyrophosphorylase
CMP-sialate synthase
cytidine 5'-monophosphosialic acid synthetase
CMP-Neu5Ac synthetase
CMP-NeuAc synthetase
acylneuraminate cytidyltransferase
CMP-N-acetylneuraminate synthetase
CMP-N-acetylneuraminate synthase
CMP-N-acetylneuraminic acid synthase
CMP-NANA synthetase
CMP-sialate synthetase
CMP-sialic synthetase
cytidine 5'-monophospho-N-acetylneuraminic acid synthetase
cytidine 5-monophosphate N-acetylneuraminic acid synthetase
cytidine monophosphosialic acid synthetase
cytidine monophosphoacetylneuraminic synthetase
cytidine monophosphosialate pyrophosphorylase
cytidine monophosphosialate synthetase
acetylneuraminate cytidylyltransferase
SynonymsEC 2.7.7.43
CMP-N-acetylneuraminic acid synthetase
CMP-NeuNAc synthetase
CMP-sialic acid synthetase
PfamPF02348 (CTP_transf_3)
[Graphical view]

KEGG pathways
MAP codePathways
MAP00530Aminosugars metabolism

UniProtKB:Accession NumberP0A0Z8
Entry nameNEUA_NEIME
ActivityCTP + N-acylneuraminate = diphosphate + CMP-N- acylneuraminate.
Subunit
Subcellular locationCytoplasm.
Cofactor

Compound table: links to PDB-related databases & PoSSuM

CofactorsSubstratesProducts
KEGG-idC00305C00063C00591C00013C01064
CompoundMagnesiumCTPN-AcylneuraminatePyrophosphateCMP-N-acylneuraminate
Typedivalent metal (Ca2+, Mg2+)amine group,nucleotideamide group,carbohydrate,carboxyl groupphosphate group/phosphate ionamide group,amine group,carbohydrate,carboxyl group,nucleotide
ChEBI18420
17677

29888

PubChem888
6176

21961011
1023

             
1eyrAUnboundAnalogue:CDPUnboundUnboundUnbound
1eyrBUnboundAnalogue:CDPUnboundUnboundUnbound
1eziAUnboundUnboundUnboundUnboundUnbound
1eziBUnboundUnboundUnboundUnboundUnbound

Active-site residues
resource
literature [9], [10]
pdbCatalytic residuesCofactor-binding residues
          
1eyrAARG 12;LYS 21;ASP 211
ASP 209;ASP 211(magnesium binding)
1eyrBARG 12;LYS 21;ASP 211
ASP 209;ASP 211(magnesium binding)
1eziAARG 12;LYS 21;ASP 211
ASP 209;ASP 211(magnesium binding)
1eziBARG 12;LYS 21;ASP 211
ASP 209;ASP 211(magnesium binding)

References for Catalytic Mechanism
ReferencesSectionsNo. of steps in catalysis
[5]Fig.5, p.7791
[9]p.8195-8196
[10]Fig.9, p.152-1531

references
[1]
PubMed ID4620387
JournalBiochem Soc Symp
Year1974
Volume(40)
Pages87-116
AuthorsSchauer R, Buscher HP, Casals-Stenzel J
TitleSialic acids: their analysis and enzymic modification in relation to the synthesis of submandibular-gland glycoproteins.
[2]
PubMed ID2544267
JournalCarbohydr Res
Year1989
Volume186
Pages326-34
AuthorsPetrie CR 3rd, Sharma M, Simmons OD, Korytnyk W
TitleSynthesis of analogs of N-acetylneuraminic acid and their effect on CMP-sialate synthase.
[3]
PubMed ID2559792
JournalCarbohydr Res
Year1989
Volume194
Pages49-61
AuthorsChristian R, Schreiner E, Zbiral E, Schulz G
TitleThe side-chain conformations of N-acetyl-7-,8-,9-deoxy-, and -4,7-dideoxy-neuraminic acid and their effect on the activation of CTP:N-acylneuraminic acid cytidylyltransferase.
[4]
PubMed ID2559791
JournalCarbohydr Res
Year1989
Volume194
Pagesc15-8
AuthorsZbiral E, Schreiner E, Christian R
TitleSynthesis of the 4-acetamido-4-deoxy analogue of N-acetylneuraminic acid and its behaviour towards CMP-sialate synthase.
[5]
PubMed ID1731934
JournalBiochemistry
Year1992
Volume31
Pages775-80
AuthorsAmbrose MG, Freese SJ, Reinhold MS, Warner TG, Vann WF
Title13C NMR investigation of the anomeric specificity of CMP-N-acetylneuraminic acid synthetase from Escherichia coli.
[6]
PubMed ID7858974
JournalBioorg Med Chem
Year1994
Volume2
Pages669-74
AuthorsLubineau A, Auge C, Gautheron-Le Narvor C, Ginet JC
TitleCombined chemical and enzymatic synthesis of the sialylated non reducing terminal sequence of GM1b glycolylated ganglioside, a potential human tumor marker.
[7]
PubMed ID9702777
JournalNat Biotechnol
Year1998
Volume16
Pages769-72
AuthorsGilbert M, Bayer R, Cunningham AM, DeFrees S, Gao Y, Watson DC, Young NM, Wakarchuk WW
TitleThe synthesis of sialylated oligosaccharides using a CMP-Neu5Ac synthetase/sialyltransferase fusion.
[8]
PubMed ID9702765
JournalNat Biotechnol
Year1998
Volume16
Pages720-1
AuthorsWarner TG
TitleSweet success with tethered enzyme catalysis.
[9]
CommentsX-ray crystallography
PubMed ID11113120
JournalJ Biol Chem
Year2001
Volume276
Pages8190-6
AuthorsMosimann SC, Gilbert M, Dombroswki D, To R, Wakarchuk W, Strynadka NC
TitleStructure of a sialic acid-activating synthetase, CMP-acylneuraminate synthetase in the presence and absence of CDP.
Related PDB1ezi,1eyr
[10]
CommentsCrystal structures of the homologous enzyme
PubMed ID11545592
JournalJ Mol Biol
Year2001
Volume312
Pages143-55
AuthorsJelakovic S, Schulz GE
TitleThe structure of CMP:2-keto-3-deoxy-manno-octonic acid synthetase and of its complexes with substrates and substrate analogs.
Related PDB1h7e,1h7f,1h7g,1h7h,1h7t
[11]
PubMed ID12355462
JournalBiotechnol Bioeng
Year2002
Volume80
Pages516-24
AuthorsLee SG, Lee JO, Yi JK, Kim BG
TitleProduction of cytidine 5'-monophosphate N-acetylneuraminic acid using recombinant Escherichia coli as a biocatalyst.
[12]
PubMed ID11893746
JournalJ Biol Chem
Year2002
Volume277
Pages19688-96
AuthorsMunster AK, Weinhold B, Gotza B, Muhlenhoff M, Frosch M, Gerardy-Schahn R
TitleNuclear localization signal of murine CMP-Neu5Ac synthetase includes residues required for both nuclear targeting and enzymatic activity.

comments
According to the literature [9], O2 atom of the substrate, NeuAc, acts as a nucleophile, which makes an SN2 attack on the alpha-phosphate of the other substrate, CTP. During the catalysis, the leaving group, pyrophsophate, can be stabilized by Arg12, Lys21, and Mg2+, which seems to bridging the phosphoryl groups of CTP and the conserved residues, Asp209 and Asp211.
The literature [9] suggested that, although there is no candidate for the general base, which can activate the nucleophile, NeuAc O2 atom, an ordered water molecule might serve as the general base or a conformational rearrangement might bring a general base within hydrogen-bonding distance of the O2 atom upon the substrate. Meanwhile, another paper [10] on its homologous enzyme, CMP-Kdo synthease, suggested that a water molecule tightly bound to two aspartic acid residues, Asp98 and Asp225, would acts as the general base, which can activate the acceptor hydroxyl group. In fact, Asp211 of this enzyme, corresponds to Asp225 of the homologous enzyme, in terms of the relative position against the nucleotide substrate. Thus, Asp225 may play a role as a general base, which can activate the acceptor O2 atom, through a water molecule.
Moreover, the paper [9] also suggested that an ordered water molecule hydrogen-bonding to mainchain oxygen atom of Asn14 can donate a proton to the leaving group, pyrophosphate.
Taken together, the reaction proceeds as follows:
(1) Asp211 acts as a general base to activate O2 hydroxyl group, through water molecule.
(2) The activated O2 atom makes a nucleophilic attack on the alpha-phosphate of CTP. The reaction proceeds via SN2-like mechanism.
(3) Arg12 and Lys21 stabilize the leaving pyrophosphate, along with magnesium ion.

createdupdated
2003-07-172009-02-26


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