MetaCyc Enzyme: purine nucleoside phosphoramidase

Gene: hinT Accession Numbers: EG12172 (MetaCyc), b1103, ECK1089

Synonyms: ycfF

Species: Escherichia coli K-12 substr. MG1655

Subunit composition of purine nucleoside phosphoramidase = [HinT]2
         purine nucleoside phosphoramidase monomer = HinT

HinT is a purine nucleoside phosphoramidase with a preference for purine nucleoside phosphoramidates. The enzyme is a homolog of the histidine triad nucleotide-binding proteins (Hint) [Chou05]. The presence of a phosphoramidase activity in E. coli was first shown by [Smith59].

In search of the physiological substrate for HinT, it was found that HinT can be adenylated by lysyl-tRNA synthetase (LysRS) in the presence of Mg2+ and lysine; the reaction is inhibited by Zn2+, Ap4A and Ap3A and enhanced by the presence of pyrophosphatase. These experiments suggest that the lysyl-AMP intermediate that is formed by LysRS is the natural substrate for HinT [Chou07].

The substrate specificity of the enzyme has been investigated; in contrast with the human enzyme, no stereochemical preference or preference for substitution at the α carboxylate was found for the E. coli enzyme [Chou07a]. The divergent C-terminal loop region of the enzyme appears to be responsible for substrate specificity [Chou07b].

hinT is essential for growth under high salt conditions [Chou05].

HinT: "histidine triad nucleotide-binding motif"

Locations: cytosol

Map Position: [1,161,108 -> 1,161,467]

Molecular Weight of Polypeptide: 13.241 kD (from nucleotide sequence)

Molecular Weight of Multimer: 27.4 kD (experimental) [Chou05]

Unification Links: ASAP:ABE-0003729 , DIP:DIP-48009N , EchoBASE:EB2090 , EcoGene:EG12172 , EcoliWiki:b1103 , Mint:MINT-1266101 , ModBase:P0ACE7 , OU-Microarray:b1103 , PortEco:hinT , PR:PRO_000022888 , Pride:P0ACE7 , Protein Model Portal:P0ACE7 , RefSeq:NP_415621 , RegulonDB:EG12172 , SMR:P0ACE7 , String:511145.b1103 , Swiss-Model:P0ACE7 , UniProt:P0ACE7

Relationship Links: InterPro:IN-FAMILY:IPR001310 , InterPro:IN-FAMILY:IPR011146 , InterPro:IN-FAMILY:IPR019808 , Panther:IN-FAMILY:PTHR23089 , PDB:Structure:3N1S , PDB:Structure:3N1T , Pfam:IN-FAMILY:PF01230 , Prints:IN-FAMILY:PR00332 , Prosite:IN-FAMILY:PS00892 , Prosite:IN-FAMILY:PS51084

Gene-Reaction Schematic: ?

Gene-Reaction Schematic

GO Terms:

Biological Process: GO:0055130 - D-alanine catabolic process Inferred from experiment [Bardaweel11]
Molecular Function: GO:0043530 - adenosine 5'-monophosphoramidase activity Inferred from experiment [Chou05]
GO:0000166 - nucleotide binding Inferred by computational analysis [UniProtGOA11]
GO:0003824 - catalytic activity Inferred by computational analysis [GOA01]
GO:0016787 - hydrolase activity Inferred by computational analysis [UniProtGOA11]
Cellular Component: GO:0005829 - cytosol Inferred from experiment Inferred by computational analysis [DiazMejia09, Ishihama08]

MultiFun Terms: metabolism

Created in EcoCyc 11-Dec-2008 by Keseler I , SRI International
Imported from EcoCyc 02-Jun-2015 by Paley S , SRI International

Enzymatic reaction of: purine nucleoside phosphoramidase

EC Number: 3.9.1.-

GMP-N-ε-(N-α-acetyl lysine methyl ester) 5'-phosphoramidate + H2O <=> GMP + N-α-acetyl lysine methyl ester

The reaction direction shown, that is, A + B ↔ C + D versus C + D ↔ A + B, is in accordance with the direction of enzyme catalysis.

The reaction is physiologically favored in the direction shown.

Alternative Substrates for GMP-N-ε-(N-α-acetyl lysine methyl ester) 5'-phosphoramidate: adenosine 5'-phosphoramidate [Chou05 ] , AMP-lysine [Chou05 ] , GMP-MOR [Chou05 ]

Imported from EcoCyc 02-Jun-2015 by Paley S , SRI International

Kinetic parameters for a number of substrates have been measured [Chou07a].

Inhibitors (Noncompetitive): TpGc (Ki = 42µM) [Bardaweel11]

Sequence Features

Feature Class Location Attached Group Citations Comment
Conserved-Region 6 -> 115  
UniProt: HIT;
Nucleotide-Phosphate-Binding-Region 30 -> 32 purine nucleotide phosporamidate
UniProt: Purine nucleotide phosphoramidate.
Amino-Acid-Sites-That-Bind 88  
UniProt: Purine nucleotide phosphoramidate.
Nucleotide-Phosphate-Binding-Region 96 -> 97 purine nucleotide phosporamidate
UniProt: Purine nucleotide phosphoramidate.
Protein-Segment 99 -> 103  
UniProt: Histidine triad motif; Sequence Annotation Type: short sequence motif.
Nucleotide-Phosphate-Binding-Region 101 -> 103 purine nucleotide phosporamidate
UniProt: Purine nucleotide phosphoramidate.
Mutagenesis-Variant 101  
[UniProt13, Bardaweel10, Bardaweel11, Chou05, UniProt13]
A or G: Abolishes enzyme activity.
Active-Site 101  
[Bardaweel10, Chou05, UniProt13]
UniProt: Tele-AMP-histidine intermediate.
Mutagenesis-Variant 114 -> 119  
[Bardaweel10, UniProt13]
UniProt: Strongly reduces enzyme activity.
Mutagenesis-Variant 117 -> 119  
[Bardaweel10, UniProt13]
UniProt: Abolishes enzyme activity.

10/20/97 Gene b1103 from Blattner lab Genbank (v. M52) entry merged into EcoCyc gene EG12172.


Bardaweel10: Bardaweel S, Pace J, Chou TF, Cody V, Wagner CR (2010). "Probing the impact of the echinT C-terminal domain on structure and catalysis." J Mol Biol 404(4);627-38. PMID: 20934431

Bardaweel11: Bardaweel S, Ghosh B, Chou TF, Sadowsky MJ, Wagner CR (2011). "E. coli histidine triad nucleotide binding protein 1 (ecHinT) is a catalytic regulator of D-alanine dehydrogenase (DadA) activity in vivo." PLoS One 6(7);e20897. PMID: 21754980

Chou05: Chou TF, Bieganowski P, Shilinski K, Cheng J, Brenner C, Wagner CR (2005). "31P NMR and genetic analysis establish hinT as the only Escherchia coli purine nucleoside phosphoramidase and as essential for growth under high salt conditions." J Biol Chem 280(15);15356-61. PMID: 15703176

Chou07: Chou TF, Wagner CR (2007). "Lysyl-tRNA synthetase generated lysyl-adenylate is a substrate for histidine triad nucleotide binding proteins." J Biol Chem 282(7):4719-27. PMID: 17158446

Chou07a: Chou TF, Baraniak J, Kaczmarek R, Zhou X, Cheng J, Ghosh B, Wagner CR (2007). "Phosphoramidate pronucleotides: a comparison of the phosphoramidase substrate specificity of human and Escherichia coli histidine triad nucleotide binding proteins." Mol Pharm 4(2);208-17. PMID: 17217311

Chou07b: Chou TF, Sham YY, Wagner CR (2007). "Impact of the C-terminal loop of histidine triad nucleotide binding protein1 (Hint1) on substrate specificity." Biochemistry 46(45);13074-9. PMID: 17939685

DiazMejia09: Diaz-Mejia JJ, Babu M, Emili A (2009). "Computational and experimental approaches to chart the Escherichia coli cell-envelope-associated proteome and interactome." FEMS Microbiol Rev 33(1);66-97. PMID: 19054114

GOA01: GOA, DDB, FB, MGI, ZFIN (2001). "Gene Ontology annotation through association of InterPro records with GO terms."

Ishihama08: Ishihama Y, Schmidt T, Rappsilber J, Mann M, Hartl FU, Kerner MJ, Frishman D (2008). "Protein abundance profiling of the Escherichia coli cytosol." BMC Genomics 9;102. PMID: 18304323

Smith59: Smith RA, Burrow DJ (1959). "Enzymic cleavage of phosphoramidic acid." Biochim Biophys Acta 34;274-6. PMID: 13832126

UniProt09: UniProt Consortium (2009). "UniProt version 15.8 released on 2009-10-01 00:00:00." Database.

UniProt13: UniProt Consortium (2013). "UniProt version 2013-08 released on 2013-08-01 00:00:00." Database.

UniProt15: UniProt Consortium (2015). "UniProt version 2015-01 released on 2015-01-16 00:00:00." Database.

UniProtGOA11: UniProt-GOA (2011). "Gene Ontology annotation based on manual assignment of UniProtKB keywords in UniProtKB/Swiss-Prot entries."

Report Errors or Provide Feedback
Please cite the following article in publications resulting from the use of MetaCyc: Caspi et al, Nucleic Acids Research 42:D459-D471 2014
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