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1oqm
From Proteopedia
(New page: 200px<br /><applet load="1oqm" size="450" color="white" frame="true" align="right" spinBox="true" caption="1oqm, resolution 2.10Å" /> '''A 1:1 complex betwee...) |
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| - | [[Image:1oqm.gif|left|200px]]<br /><applet load="1oqm" size=" | + | [[Image:1oqm.gif|left|200px]]<br /><applet load="1oqm" size="350" color="white" frame="true" align="right" spinBox="true" |
caption="1oqm, resolution 2.10Å" /> | caption="1oqm, resolution 2.10Å" /> | ||
'''A 1:1 complex between alpha-lactalbumin and beta1,4-galactosyltransferase in the presence of UDP-N-acetyl-galactosamine'''<br /> | '''A 1:1 complex between alpha-lactalbumin and beta1,4-galactosyltransferase in the presence of UDP-N-acetyl-galactosamine'''<br /> | ||
==Overview== | ==Overview== | ||
| - | beta1,4-Galactosyltransferase I (Gal-T1) normally transfers Gal from | + | beta1,4-Galactosyltransferase I (Gal-T1) normally transfers Gal from UDP-Gal to GlcNAc in the presence of Mn(2+) ion. In the presence of alpha-lactalbumin (LA), the Gal acceptor specificity is altered from GlcNAc to Glc. Gal-T1 also transfers GalNAc from UDP-GalNAc to GlcNAc, but with only approximately 0.1% of Gal-T activity. To understand this low GalNAc-transferase activity, we have carried out the crystal structure analysis of the Gal-T1.LA complex with UDP-GalNAc at 2.1-A resolution. The crystal structure reveals that the UDP-GalNAc binding to Gal-T1 is similar to the binding of UDP-Gal to Gal-T1, except for an additional hydrogen bond formed between the N-acetyl group of GalNAc moiety with the Tyr-289 side chain hydroxyl group. Elimination of this additional hydrogen bond by mutating Tyr-289 residue to Leu, Ile, or Asn enhances the GalNAc-transferase activity. Although all three mutants exhibit enhanced GalNAc-transferase activity, the mutant Y289L exhibits GalNAc-transferase activity that is nearly 100% of its Gal-T activity, even while completely retaining its Gal-T activity. The steady state kinetic analyses on the Leu-289 mutant indicate that the K(m) for GlcNAc has increased compared to the wild type. On the other hand, the catalytic constant (k(cat)) in the Gal-T reaction is comparable with the wild type, whereas it is 3-5-fold higher in the GalNAc-T reaction. Interestingly, in the presence of LA, these mutants also transfer GalNAc to Glc instead of to GlcNAc. The present study demonstrates that, in the Gal-T family, the Tyr-289/Phe-289 residue largely determines the sugar donor specificity. |
==About this Structure== | ==About this Structure== | ||
| - | 1OQM is a [http://en.wikipedia.org/wiki/Protein_complex Protein complex] structure of sequences from [http://en.wikipedia.org/wiki/Bos_taurus Bos taurus] and [http://en.wikipedia.org/wiki/Mus_musculus Mus musculus] with CA, MN, UD2 and PG4 as [http://en.wikipedia.org/wiki/ligands ligands]. This structure | + | 1OQM is a [http://en.wikipedia.org/wiki/Protein_complex Protein complex] structure of sequences from [http://en.wikipedia.org/wiki/Bos_taurus Bos taurus] and [http://en.wikipedia.org/wiki/Mus_musculus Mus musculus] with <scene name='pdbligand=CA:'>CA</scene>, <scene name='pdbligand=MN:'>MN</scene>, <scene name='pdbligand=UD2:'>UD2</scene> and <scene name='pdbligand=PG4:'>PG4</scene> as [http://en.wikipedia.org/wiki/ligands ligands]. This structure supersedes the now removed PDB entry 1L7W. Active as [http://en.wikipedia.org/wiki/Beta-N-acetylglucosaminylglycopeptide_beta-1,4-galactosyltransferase Beta-N-acetylglucosaminylglycopeptide beta-1,4-galactosyltransferase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=2.4.1.38 2.4.1.38] Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1OQM OCA]. |
==Reference== | ==Reference== | ||
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[[Category: Mus musculus]] | [[Category: Mus musculus]] | ||
[[Category: Protein complex]] | [[Category: Protein complex]] | ||
| - | [[Category: Qasba, P | + | [[Category: Qasba, P K.]] |
[[Category: Ramakrishnan, B.]] | [[Category: Ramakrishnan, B.]] | ||
[[Category: CA]] | [[Category: CA]] | ||
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[[Category: udp-galnac]] | [[Category: udp-galnac]] | ||
| - | ''Page seeded by [http:// | + | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Feb 21 14:20:30 2008'' |
Revision as of 12:20, 21 February 2008
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A 1:1 complex between alpha-lactalbumin and beta1,4-galactosyltransferase in the presence of UDP-N-acetyl-galactosamine
Overview
beta1,4-Galactosyltransferase I (Gal-T1) normally transfers Gal from UDP-Gal to GlcNAc in the presence of Mn(2+) ion. In the presence of alpha-lactalbumin (LA), the Gal acceptor specificity is altered from GlcNAc to Glc. Gal-T1 also transfers GalNAc from UDP-GalNAc to GlcNAc, but with only approximately 0.1% of Gal-T activity. To understand this low GalNAc-transferase activity, we have carried out the crystal structure analysis of the Gal-T1.LA complex with UDP-GalNAc at 2.1-A resolution. The crystal structure reveals that the UDP-GalNAc binding to Gal-T1 is similar to the binding of UDP-Gal to Gal-T1, except for an additional hydrogen bond formed between the N-acetyl group of GalNAc moiety with the Tyr-289 side chain hydroxyl group. Elimination of this additional hydrogen bond by mutating Tyr-289 residue to Leu, Ile, or Asn enhances the GalNAc-transferase activity. Although all three mutants exhibit enhanced GalNAc-transferase activity, the mutant Y289L exhibits GalNAc-transferase activity that is nearly 100% of its Gal-T activity, even while completely retaining its Gal-T activity. The steady state kinetic analyses on the Leu-289 mutant indicate that the K(m) for GlcNAc has increased compared to the wild type. On the other hand, the catalytic constant (k(cat)) in the Gal-T reaction is comparable with the wild type, whereas it is 3-5-fold higher in the GalNAc-T reaction. Interestingly, in the presence of LA, these mutants also transfer GalNAc to Glc instead of to GlcNAc. The present study demonstrates that, in the Gal-T family, the Tyr-289/Phe-289 residue largely determines the sugar donor specificity.
About this Structure
1OQM is a Protein complex structure of sequences from Bos taurus and Mus musculus with , , and as ligands. This structure supersedes the now removed PDB entry 1L7W. Active as Beta-N-acetylglucosaminylglycopeptide beta-1,4-galactosyltransferase, with EC number 2.4.1.38 Full crystallographic information is available from OCA.
Reference
Structure-based design of beta 1,4-galactosyltransferase I (beta 4Gal-T1) with equally efficient N-acetylgalactosaminyltransferase activity: point mutation broadens beta 4Gal-T1 donor specificity., Ramakrishnan B, Qasba PK, J Biol Chem. 2002 Jun 7;277(23):20833-9. Epub 2002 Mar 26. PMID:11916963
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