Tetrahedron Letters
Novel preloaded resins for solid-phase biotinylation of carboxylic acids
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Nadezda Cankarova, Petr Funk, Jan Hlavac, Miroslav Soural
Department of Organic Chemistry, Institute of Molecular and Translational Medicine, Faculty of Science, University of Palacky, 17. listopadu 12, 771 46 Olomouc, Czech Republic
a r t i c l e i n f o
a b s t r a c t
Article history:
Use of solid-phase synthesis for the derivatization of carboxylic acids with biotinylated spacers consisting
of ethylenoxy units is described. An aminomethylated resin provided with an acid-labile aldehyde linker
is used as the polymer support and three different systems with a reactive amino group are introduced.
Acylation of each system was tested with a set of model carboxylic acids and afforded crude products of
excellent purity. The preloaded resins are similar to the Biotin-PEG-NovaTagTM resin but offer several
advantages including simple elongation of the spacer arm. The protocols described represent a very effi-
cient way of modifying compounds to obtain ligands for affinity chromatography studies.
Ó 2011 Elsevier Ltd. All rights reserved.
Received 29 May 2011
Revised 9 August 2011
Accepted 19 August 2011
Available online 26 August 2011
Keywords:
Solid-phase synthesis
Biotine
PEG linker
Acylation
Affinity chromatography
Biotinylation of organic compounds is a very frequent approach
for detailed studies of biochemical properties of biologically active
substances. Biotinylated agents are widely used to immobilize
compounds for affinity chromatography,1,2 labeling of peptides,3,4
proteins,5–7 nucleic acids, antibodies,8 etc. When the molecule to
be studied is suitably derivatized with biotin, it can be simply
immobilized on a chromatography column by using its highly spe-
cific and tight non-covalent interaction with avidin or streptavi-
din.9 When the selected compound is modified for an affinity
chromatography assay, the key task is incorporation of the spacer
between the molecule to be studied and biotin.10–12 The spacer
must meet the following basic criteria: (i) it has to be long enough
to avoid steric repulsion between avidin and the target biomole-
cule which interacts with the immobilized compound, (ii) it has
to be inert to non-specific interactions with biomolecules that do
not interact with the immobilized compound, (iii) it has to avoid
decreasing the biotinylated system’s solubility in water. Aliphatic
spacers (for instance, a spacer consisting of the caproic acid skele-
ton) have been used for a long time. However, more recently they
have been successfully replaced by ethylenoxy units—containing
ligands (ethylene glycols—EGs) which fulfill the last two men-
tioned criteria. For applications in which a longer spacer is required
(e.g., more than 12 atoms) EGs, in contrast to aliphatic chains,
avoid hydrophobic interactions resulting in deformation of the
spacer.
with carboxy group-containing compounds, (ii) acylation of an
amino group-containing compound with the biotin-EGs-COOH
system. In this Letter we describe an efficient method of biotinyla-
tion of carboxylic acids using the first mentioned method. So far,
biotinylation of appropriate compounds via EG linkers has been
commonly performed using solution-phase synthesis and/or
commercially available biotin-EG-NH2 systems. However, solution-
phase modification of compounds is usually complicated by diffi-
cult and time-consuming isolation of products from reaction
mixtures and also complicated purification of intermediates as well
as of the final products. Such problems can be eliminated when the
concept of solid-phase synthesis is applied. There are two alterna-
tives to the use of solid-phase synthesis for biotinylation of organic
compounds: (i) the substrate which undergoes biotinylation is pre-
immobilized (or directly synthesized) on an insoluble polymer
support.13 (ii) the biotin-EG-NH2 system is immobilized and the
resulting preloaded resin is used for an acylation reaction with
an appropriate carboxy group-containing compound. Quite re-
cently, Novabiochem has introduced the first preloaded resin of
this kind: Biotin-PEG-NovaTag™ resin, which has already been
successfully used in practice in several cases.14–16 In this Letter,
we describe preparation and use of alternative systems which offer
O
O
O
The most frequent way to attach the compound to be studied to
the biotin-EG system is: (i) acylation of the biotin-EG-NH2 system
HO
O
O
O HOBt, DIC
H
NH2
N
L
3O
O
DCM/DMF (1:1)
rt, overnight
O
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Corresponding author. Tel.: +420 585634418; fax: +420 585634465.
Scheme 1. Attachment of an acid-labile linker to the aminomethylated resin.
0040-4039/$ - see front matter Ó 2011 Elsevier Ltd. All rights reserved.