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Structural effects of naturally occurring human blood group B galactosyltransferase mutations adjacent to the DXD motif

Persson, Mattias; Letts, James A.; Hosseini Maaf, Bahram LU ; Borisova, Svetlana N.; Palcic, Monica M.; Evans, Stephen V. and Olsson, Martin L LU (2007) In Journal of Biological Chemistry 282(13). p.9564-9570
Abstract
Human blood group A and B antigens are produced by two closely related glycosyltransferase enzymes. An N-acetylgalactosaminyltransferase (GTA) utilizes UDP-GalNAc to extend H antigen acceptors (Fuc alpha(1-2)Gal beta-OR) producing A antigens, whereas a galactosyltransferase (GTB) utilizes UDP-Gal as a donor to extend H structures producing B antigens. GTA and GTB have a characteristic (DVD213)-D-211 motif that coordinates to a Mn2+ ion shown to be critical in donor binding and catalysis. Three GTB mutants, M214V, M214T, and M214R, with alterations adjacent to the (211)DVD213 motif have been identified in blood banking laboratories. From serological phenotyping, individuals with the M214R mutation show the B,1 variant expressing very low... (More)
Human blood group A and B antigens are produced by two closely related glycosyltransferase enzymes. An N-acetylgalactosaminyltransferase (GTA) utilizes UDP-GalNAc to extend H antigen acceptors (Fuc alpha(1-2)Gal beta-OR) producing A antigens, whereas a galactosyltransferase (GTB) utilizes UDP-Gal as a donor to extend H structures producing B antigens. GTA and GTB have a characteristic (DVD213)-D-211 motif that coordinates to a Mn2+ ion shown to be critical in donor binding and catalysis. Three GTB mutants, M214V, M214T, and M214R, with alterations adjacent to the (211)DVD213 motif have been identified in blood banking laboratories. From serological phenotyping, individuals with the M214R mutation show the B,1 variant expressing very low levels of B antigens, whereas those with M214T and M214V mutations give rise to A(weak)B phenotypes. Kinetic analysis of recombinant mutant GTB enzymes revealed that M214R has a 1200-fold decrease in k(cat) compared with wild type GTB. The crystal structure of M214R showed that DVD motif coordination to Mn2+ was disrupted by Arg-214 causing displacement of the metal by a water molecule. Kinetic characterizations of the M214T and M214V mutants revealed they both had GTA and GTB activity consistent with the serology. The crystal structure of the M214T mutant showed no change in DVD coordination to Mn2+. Instead a critical residue, Met-266, which is responsible for determining donor specificity, had adopted alternate conformations. The conformation with the highest occupancy opens up the active site to accommodate the larger A-specific donor, UDP-GalNAc, accounting for the dual specificity. (Less)
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author
organization
publishing date
type
Contribution to journal
publication status
published
subject
in
Journal of Biological Chemistry
volume
282
issue
13
pages
9564 - 9570
publisher
ASBMB
external identifiers
  • wos:000245421700029
  • scopus:34248229370
ISSN
1083-351X
DOI
10.1074/jbc.M610998200
language
English
LU publication?
yes
id
3685dec4-19f3-4fc0-a0bd-11d52d0846c2 (old id 667524)
date added to LUP
2007-12-18 14:50:20
date last changed
2017-10-22 03:44:51
@article{3685dec4-19f3-4fc0-a0bd-11d52d0846c2,
  abstract     = {Human blood group A and B antigens are produced by two closely related glycosyltransferase enzymes. An N-acetylgalactosaminyltransferase (GTA) utilizes UDP-GalNAc to extend H antigen acceptors (Fuc alpha(1-2)Gal beta-OR) producing A antigens, whereas a galactosyltransferase (GTB) utilizes UDP-Gal as a donor to extend H structures producing B antigens. GTA and GTB have a characteristic (DVD213)-D-211 motif that coordinates to a Mn2+ ion shown to be critical in donor binding and catalysis. Three GTB mutants, M214V, M214T, and M214R, with alterations adjacent to the (211)DVD213 motif have been identified in blood banking laboratories. From serological phenotyping, individuals with the M214R mutation show the B,1 variant expressing very low levels of B antigens, whereas those with M214T and M214V mutations give rise to A(weak)B phenotypes. Kinetic analysis of recombinant mutant GTB enzymes revealed that M214R has a 1200-fold decrease in k(cat) compared with wild type GTB. The crystal structure of M214R showed that DVD motif coordination to Mn2+ was disrupted by Arg-214 causing displacement of the metal by a water molecule. Kinetic characterizations of the M214T and M214V mutants revealed they both had GTA and GTB activity consistent with the serology. The crystal structure of the M214T mutant showed no change in DVD coordination to Mn2+. Instead a critical residue, Met-266, which is responsible for determining donor specificity, had adopted alternate conformations. The conformation with the highest occupancy opens up the active site to accommodate the larger A-specific donor, UDP-GalNAc, accounting for the dual specificity.},
  author       = {Persson, Mattias and Letts, James A. and Hosseini Maaf, Bahram and Borisova, Svetlana N. and Palcic, Monica M. and Evans, Stephen V. and Olsson, Martin L},
  issn         = {1083-351X},
  language     = {eng},
  number       = {13},
  pages        = {9564--9570},
  publisher    = {ASBMB},
  series       = {Journal of Biological Chemistry},
  title        = {Structural effects of naturally occurring human blood group B galactosyltransferase mutations adjacent to the DXD motif},
  url          = {http://dx.doi.org/10.1074/jbc.M610998200},
  volume       = {282},
  year         = {2007},
}