C O M M U N I C A T I O N S
Scheme 6. Synthesis of Lipidated Pseudodisaccharide 3a
to afford the desired R-pseudopentasaccharide in 64% yield. Overnight
treatment of the product with Et3N·3HF removed the TBS and TES
groups to provide diol 36, of which the anomeric JCH coupling constants
confirmed R stereochemistry for all mannose units.14 Installation of
the phosphoethanolamine group by treatment with phosphoramidite 4
for a short period (1 h) was selective for the Man-III 6-O-position to
give 37.15 To obtain the target GPI 1, a three-step, one-pot deprotection
protocol was developed to efficiently remove all of the protecting
groups from 37 in 4 h: (i) Zn-mediated reduction of the azide; (ii)
removal of the base-labile Fmoc and cyanoethoxy groups with DBU;
(iii) hydrolysis of all PMB ethers with 10% TFA. The target GPI
anchor 1 was finally obtained in 81% yield after purification with a
Sephadex LH-20 column and was characterized with 1H NMR
spectroscopy and MALDI-TOF MS.
a Conditions: (a) TMSOTf (cat.), MS 4 Å, Et2O, 44%. (b) [Ir(COD)-
(PMePh2)2]PF6, H2, THF; then HgCl2, HgO, acetone, H2O, 96%. (c) 10,
tetrazole, CH3CN, CH2Cl2; then tBuOOH, -40 °C. (d) Et3N·3HF, THF,
CH3CN, 56% (two steps).
Scheme 7. Assembly of GPI Anchor 1a
In summary, a GPI anchor containing unsaturated lipid chains
was efficiently synthesized using the PMB group for hydroxyl
protection. This represents a potentially generally useful strategy
for the design and synthesis of uniquely functionalized GPIs, as
well as other complex oligosaccharides, which may not be easily
accessible by means of traditional protection methods, such as using
benzyl, acetyl, or benzoyl groups. We are currently using this
synthetic strategy to prepare various biofunctionalized GPI anchors
aimed at probing cell surface GPIomics.
Acknowledgment. This work was funded by NIH/NIGMS
(R01GM090270). We thank Dr. B. Shay and Dr. L. Hryhorczuk
for MS measurements, and Dr. B. Ksebati for help with some NMR
spectroscopy experiments.
Supporting Information Available: Experimental procedures and
spectroscopic data. This material is available free of charge via the
References
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°C, 50% for 37 (61% BRSM). (d) Zn, AcOH, CH2Cl2, 2 h; DBU, CH2Cl2
1 h; CH2Cl2-TFA (9:1), 1 h, 81% (three steps).
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pseudodisaccharide 34 (Scheme 6). While the conversion rate was very
good (84%), stereoselectivity slightly favored the undesired ꢀ anomer
(R/ꢀ 1.0:1.6). However, when Et2O was used as the solvent, a
moderately improved stereoselectivity was obtained (R/ꢀ 1.2:1.0). After
the anomeric mixture was separated by preparative HPLC (separation
by silica gel column following the next step was also possible), the
Ir/Hg deallylation protocol was utilized to expose the inositol 1-O-
position, giving 35 in 96% yield. Next, the unsaturated phospholipid
was installed by reaction with freshly prepared phosphoramidite 10
under the influence of tetrazole. The intermediate phosphite was
selectively oxidized in situ to a phosphate using tert-butyl hydroper-
oxide at -40 °C. Exposure of the product to Et3N·3HF for 5 days
removed the presumably hindered 4-O-TBS group, which gave
compound 3 as a 1:1 diastereomeric mixture, originating from the
stereogenic phosphorus atom, in 56% yield over two steps. The
resulting mixture was separated by preparative HPLC to facilitate the
characterization of 3 and subsequent complex intermediates.
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(11) The correct enantiomer was identified by comparison of optical rotation
with an authentic sample (see Supporting Information).
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(14) See Supporting Information.
(15) Regiochemistry of 37 was confirmed by 1H/COSY NMR spectroscopy:
The Man-I 2-H signal did not shift downfield, as would be expected if
phosphorylation occurred at this site.
The key step in the final stage of the synthesis was to couple the
trimannose and pseudodisaccharide fragments (Scheme 7). Trimannose
imidate 2 reacted smoothly with 3 in the presence of catalytic TMSOTf
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