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DOI no:
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EMBO J
27:234-243
(2008)
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PubMed id:
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Structure-guided identification of a new catalytic motif of oligosaccharyltransferase.
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M.Igura,
N.Maita,
J.Kamishikiryo,
M.Yamada,
T.Obita,
K.Maenaka,
D.Kohda.
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ABSTRACT
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Asn-glycosylation is widespread not only in eukaryotes but also in archaea and
some eubacteria. Oligosaccharyltransferase (OST) catalyzes the co-translational
transfer of an oligosaccharide from a lipid donor to an asparagine residue in
nascent polypeptide chains. Here, we report that a thermophilic archaeon,
Pyrococcus furiosus OST is composed of the STT3 protein alone, and catalyzes the
transfer of a heptasaccharide, containing one hexouronate and two pentose
residues, onto peptides in an Asn-X-Thr/Ser-motif-dependent manner. We also
determined the 2.7-A resolution crystal structure of the C-terminal soluble
domain of Pyrococcus STT3. The structure-based multiple sequence alignment
revealed a new motif, DxxK, which is adjacent to the well-conserved WWDYG motif
in the tertiary structure. The mutagenesis of the DK motif residues in yeast
STT3 revealed the essential role of the motif in the catalytic activity. The
function of this motif may be related to the binding of the pyrophosphate group
of lipid-linked oligosaccharide donors through a transiently bound cation. Our
structure provides the first structural insights into the formation of the
oligosaccharide-asparagine bond.
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Selected figure(s)
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Figure 3.
Figure 3 Crystal structure of the C-terminal soluble domain of
STT3. (A) Domain structure of P. furiosus STT3. TM,
transmembrane domain; CC, central core domain, residues
471–600+683–725; IS, insertion domain, residues 601–682;
P1, peripheral domain 1, residues 726–821; P2, peripheral
domain 2, residues 822–967. The position of the WWDYG motif is
indicated by an asterisk. (B) Stereoview of the overall
structure of the C-terminal soluble domain of STT3 (residues
471–967). The WWDYG motif is shown in magenta. The disulfide
bond between C638 and C658 is shown as yellow sticks. A bound
metal cation is shown as a yellow sphere. (C) Different view
from (B).
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Figure 4.
Figure 4 Putative active site of oligosaccharyltransferase
comprising the two conserved motifs. (A) Sequence alignment of
the region containing the known WWDYG motif and the newly found
DK motif. An initial alignment was obtained with the Mafft
algorithm (Katoh et al, 2005) in the program Jalview, and then
was edited manually. (B) Close-up view of the putative active
site, with the WWDYG motif in cyan (W511, W512, D513, Y514, and
G515), and the DK motif in yellow (D571 and K574). Alternative
possible side-chain directions of W512 and D513 in the absence
of crystal packing effects are indicated by gray arrows.
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The above figures are
reprinted
by permission from Macmillan Publishers Ltd:
EMBO J
(2008,
27,
234-243)
copyright 2008.
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Figures were
selected
by the author.
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Literature references that cite this PDB file's
key reference
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PubMed id
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Reference
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A.Vik,
F.E.Aas,
J.H.Anonsen,
S.Bilsborough,
A.Schneider,
W.Egge-Jacobsen,
and
M.Koomey
(2009).
Broad spectrum O-linked protein glycosylation in the human pathogen Neisseria gonorrhoeae.
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Proc Natl Acad Sci U S A, 106,
4447-4452.
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B.Chaban,
S.M.Logan,
J.F.Kelly,
and
K.F.Jarrell
(2009).
AglC and AglK are involved in biosynthesis and attachment of diacetylated glucuronic acid to the N-glycan in Methanococcus voltae.
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J Bacteriol, 191,
187-195.
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B.L.Schulz,
C.U.Stirnimann,
J.P.Grimshaw,
M.S.Brozzo,
F.Fritsch,
E.Mohorko,
G.Capitani,
R.Glockshuber,
M.G.Grütter,
and
M.Aebi
(2009).
Oxidoreductase activity of oligosaccharyltransferase subunits Ost3p and Ost6p defines site-specific glycosylation efficiency.
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Proc Natl Acad Sci U S A, 106,
11061-11066.
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K.Hese,
C.Otto,
F.H.Routier,
and
L.Lehle
(2009).
The yeast oligosaccharyltransferase complex can be replaced by STT3 from Leishmania major.
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Glycobiology, 19,
160-171.
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L.Izquierdo,
B.L.Schulz,
J.A.Rodrigues,
M.L.Güther,
J.B.Procter,
G.J.Barton,
M.Aebi,
and
M.A.Ferguson
(2009).
Distinct donor and acceptor specificities of Trypanosoma brucei oligosaccharyltransferases.
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EMBO J, 28,
2650-2661.
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M.Kämpf,
B.Absmanner,
M.Schwarz,
and
L.Lehle
(2009).
Biochemical Characterization and Membrane Topology of Alg2 from Saccharomyces cerevisiae as a Bifunctional {alpha}1,3- and 1,6-Mannosyltransferase Involved in Lipid-linked Oligosaccharide Biosynthesis.
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J Biol Chem, 284,
11900-11912.
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S.Yurist-Doutsch,
and
J.Eichler
(2009).
Manual annotation, transcriptional analysis, and protein expression studies reveal novel genes in the agl cluster responsible for N glycosylation in the halophilic archaeon Haloferax volcanii.
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J Bacteriol, 191,
3068-3075.
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C.M.Wilson,
Q.Roebuck,
and
S.High
(2008).
Ribophorin I regulates substrate delivery to the oligosaccharyltransferase core.
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Proc Natl Acad Sci U S A, 105,
9534-9539.
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L.L.Lairson,
B.Henrissat,
G.J.Davies,
and
S.G.Withers
(2008).
Glycosyltransferases: structures, functions, and mechanisms.
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Annu Rev Biochem, 77,
521-555.
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M.W.Vetting,
P.A.Frantom,
and
J.S.Blanchard
(2008).
Structural and enzymatic analysis of MshA from Corynebacterium glutamicum: substrate-assisted catalysis.
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J Biol Chem, 283,
15834-15844.
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PDB codes:
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N.Plavner,
and
J.Eichler
(2008).
Defining the topology of the N-glycosylation pathway in the halophilic archaeon Haloferax volcanii.
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J Bacteriol, 190,
8045-8052.
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S.Yurist-Doutsch,
B.Chaban,
D.J.VanDyke,
K.F.Jarrell,
and
J.Eichler
(2008).
Sweet to the extreme: protein glycosylation in Archaea.
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Mol Microbiol, 68,
1079-1084.
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S.Yurist-Doutsch,
M.Abu-Qarn,
F.Battaglia,
H.R.Morris,
P.G.Hitchen,
A.Dell,
and
J.Eichler
(2008).
AglF, aglG and aglI, novel members of a gene island involved in the N-glycosylation of the Haloferax volcanii S-layer glycoprotein.
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Mol Microbiol, 69,
1234-1245.
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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.
|
472 a.a.
_CL
_CA
