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PDBsum entry 1vzx

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protein ligands metals Protein-protein interface(s) links
Transferase PDB id
1vzx
Jmol
Contents
Protein chains
287 a.a. *
Ligands
GAL-NAG ×2
UDP ×2
GOL ×2
Metals
_MN ×2
Waters ×642
* Residue conservation analysis
PDB id:
1vzx
Name: Transferase
Title: Roles of active site tryptophans in substrate binding and catalysis by alpha-1,3 galactosyltransferase
Structure: N-acetyllactosaminide alpha-1,3- galactosyltransferase. Chain: a, b. Fragment: catalytic domain, residues 80-368. Synonym: galactosyltransferase, udp-galactose, beta-d-galactosyl-1,4-n-acetyl-d-glucosaminide alpha-1,3-galactosyltransferase. Engineered: yes. Mutation: yes.
Source: Bos taurus. Bovine. Organism_taxid: 9913. Expressed in: escherichia coli. Expression_system_taxid: 562.
Resolution:
1.97Å     R-factor:   0.154     R-free:   0.202
Authors: Y.Zhang,A.Deshpande,Z.Xie,R.Natesh,K.R.Acharya,K.Brew
Key ref: Y.Zhang et al. (2004). Roles of active site tryptophans in substrate binding and catalysis by alpha-1,3 galactosyltransferase. Glycobiology, 14, 1295-1302. PubMed id: 15229192 DOI: 10.1093/glycob/cwh119
Date:
28-May-04     Release date:   08-Jul-04    
PROCHECK
Go to PROCHECK summary
 Headers
 References

Protein chains
Pfam   ArchSchema ?
P14769  (GGTA1_BOVIN) -  N-acetyllactosaminide alpha-1,3-galactosyltransferase
Seq:
Struc:
368 a.a.
287 a.a.*
Key:    PfamA domain  Secondary structure  CATH domain
* PDB and UniProt seqs differ at 1 residue position (black cross)

 Enzyme reactions 
   Enzyme class: E.C.2.4.1.87  - N-acetyllactosaminide 3-alpha-galactosyltransferase.
[IntEnz]   [ExPASy]   [KEGG]   [BRENDA]
      Reaction: UDP-alpha-D-galactose + beta-D-galactosyl-(1->4)-beta-N-acetyl-D- glucosaminyl-R = UDP + alpha-D-galactosyl-(1->3)-beta-D-galactosyl- (1->4)-beta-N-acetylglucosaminyl-R
UDP-alpha-D-galactose
+
beta-D-galactosyl-(1->4)-beta-N-acetyl-D- glucosaminyl-R
Bound ligand (Het Group name = NAG)
matches with 55.00% similarity
=
UDP
Bound ligand (Het Group name = UDP)
corresponds exactly
+ alpha-D-galactosyl-(1->3)-beta-D-galactosyl- (1->4)-beta-N-acetylglucosaminyl-R
Molecule diagrams generated from .mol files obtained from the KEGG ftp site
 Gene Ontology (GO) functional annotation 
  GO annot!
  Cellular component     membrane   1 term 
  Biological process     carbohydrate metabolic process   1 term 
  Biochemical function     transferase activity, transferring hexosyl groups     1 term  

 

 
    Added reference    
 
 
DOI no: 10.1093/glycob/cwh119 Glycobiology 14:1295-1302 (2004)
PubMed id: 15229192  
 
 
Roles of active site tryptophans in substrate binding and catalysis by alpha-1,3 galactosyltransferase.
Y.Zhang, A.Deshpande, Z.Xie, R.Natesh, K.R.Acharya, K.Brew.
 
  ABSTRACT  
 
Aromatic amino acids are frequent components of the carbohydrate binding sites of lectins and enzymes. Previous structural studies have shown that in alpha-1,3 galactosyltransferase, the binding site for disaccharide acceptor substrates is encircled by four tryptophans, residues 249, 250, 314, and 356. To investigate their roles in enzyme specificity and catalysis, we expressed and characterized variants of the catalytic domain of alpha-1,3 galactosyltransferase with substitutions for each tryptophan. Substitution of glycine for tryptophan 249, whose indole ring interacts with the nonpolar B face of glucose or GlcNAc, greatly increases the K(m) for the acceptor substrate. In contrast, the substitution of tyrosine for tryptophan 314, which interacts with the beta-galactosyl moiety of the acceptor and UDP-galactose, decreases k(cat) for the galactosyltransferase reaction but does not affect the low UDP-galactose hydrolase activity. Thus, this highly conserved residue stabilizes the transition state for the galactose transfer to disaccharide but not to water. High-resolution crystallographic structures of the Trp(249)Gly mutant and the Trp(314)Tyr mutant indicate that the mutations do not affect the overall structure of the enzyme or its interactions with ligands. Substitutions for tryptophan 250 have only small effects on catalytic activity, but mutation of tryptophan 356 to threonine reduces catalytic activity for both transferase and hydrolase activities and reduces affinity for the acceptor substrate. This residue is adjacent to the flexible C-terminus that becomes ordered on binding UDP to assemble the acceptor binding site and influence catalysis. The results highlight the diverse roles of these tryptophans in enzyme action and the importance of k(cat) changes in modulating glycosyltransferase specificity.
 

Literature references that cite this PDB file's key reference

  PubMed id Reference
21057479 C.J.Bosques, B.E.Collins, J.W.Meador, H.Sarvaiya, J.L.Murphy, G.Dellorusso, D.A.Bulik, I.H.Hsu, N.Washburn, S.F.Sipsey, J.R.Myette, R.Raman, Z.Shriver, R.Sasisekharan, and G.Venkataraman (2010).
Chinese hamster ovary cells can produce galactose-α-1,3-galactose antigens on proteins.
  Nat Biotechnol, 28, 1153-1156.  
19622749 P.Tumbale, and K.Brew (2009).
Characterization of a metal-independent CAZy family 6 glycosyltransferase from Bacteroides ovatus.
  J Biol Chem, 284, 25126-25134.  
18047841 B.A.Macher, and U.Galili (2008).
The Galalpha1,3Galbeta1,4GlcNAc-R (alpha-Gal) epitope: a carbohydrate of unique evolution and clinical relevance.
  Biochim Biophys Acta, 1780, 75-88.  
18782853 P.Tumbale, H.Jamaluddin, N.Thiyagarajan, K.R.Acharya, and K.Brew (2008).
Screening a limited structure-based library identifies UDP-GalNAc-specific mutants of alpha-1,3-galactosyltransferase.
  Glycobiology, 18, 1036-1043.
PDB codes: 2vxl 2vxm
17057723 L.L.Lairson, A.G.Watts, W.W.Wakarchuk, and S.G.Withers (2006).
Using substrate engineering to harness enzymatic promiscuity and expand biological catalysis.
  Nat Chem Biol, 2, 724-728.  
16620397 M.S.Sujatha, and P.V.Balaji (2006).
Fold-recognition and comparative modeling of human alpha2,3-sialyltransferases reveal their sequence and structural similarities to CstII from Campylobacter jejuni.
  BMC Struct Biol, 6, 9.  
The most recent references are shown first. Citation data come partly from CiteXplore and partly from an automated harvesting procedure. Note that this is likely to be only a partial list as not all journals are covered by either method. However, we are continually building up the citation data so more and more references will be included with time. Where a reference describes a PDB structure, the PDB codes are shown on the right.