Reversible Covalent Bond Toolbox

Yunfeng Cheng, Hanjing Peng, and Binghe Wang

Georgia State University, Atlanta, GA, USA

1 Introduction
2 Interactions between the Boronic Acid Unit and
Nucleophiles/Lewis Bases
3 Electron-Deficient Carbonyl Groups and Their
Interactions with Hydroxyl Groups and Amino
Group
4 MetalAnion/Ligand Interactions
5 Chemosensors Based on the Michael Addition
Reaction
6 Conclusion
Acknowledgments
References

1
1

4
8
10
10
10
10

INTRODUCTION

In supramolecular chemistry, functional group interactions

play a critical role. The use of covalent interactions, however, is not as common as noncovalent interactions mostly
because of their irreversible nature. This chapter covers those covalent interactions that are readily reversible
and are often used in the construction of supramolecular
assemblies.

INTERACTIONS BETWEEN THE

BORONIC ACID UNIT AND
NUCLEOPHILES/LEWIS BASES

In applying covalent bond formation in supramolecular chemistry, boronic acids stand out as one of the

most commonly used functional groups because of their

ability to reversibly interact with nucleophiles/Lewis bases.
Scheme 1 shows the binding processes involving boronic
acid. The boron atom of boronic acid 1 has an open shell
with only six valence electrons in its trigonal neutral form,
which renders 1 a Lewis acid and endows its reactivity
with nucleophiles/Lewis bases. As a result, 1 can easily
react with a protic solvent leading to the formation of the
anionic tetrahedron form (2), accompanied with the release
of a proton, which is the origin of its Br
nsted acidity.
Besides, 1 can also react with other nucleophiles such as
cyanide, fluoride, and hydroxyl groups as well as amino
groups. As a further extension of its ability to interact with
nucleophiles, the boronic acid moiety has a unique ability to bind to compounds with two or three nucleophiles
strategically positioned to give divalent or trivalent binding. As a result, boronic acid is known to interact with
diols, -aminoacids, alcohols, aminoalcohols, -hydroxyl
acids, as well as fluoride and cyanide.14 For example,
boronic acid is considered as one of the most promising
building blocks for the design of sensors for carbohydrates
in aqueous solution, due to its intrinsic binding affinity
with multiple hydroxyl groups, which exist on most carbohydrates.4 The specific interactions involve covalent bond
formation with two neighboring hydroxyl groups. In some
cases, trivalent interactions are also possible. Such is the
case of sorbitol binding with phenylboronic acid. Because
carbohydrate recognition is the most common applications,
subsequent discussions of the use of the boronic acid moiety
are focused on carbohydrate sensing.
In designing boronic acid-based receptors for carbohydrates (and other hydroxyl containing compounds for
various applications, it is important to take into consideration the key factors including the pKa values of the boronic
acid and the diol, the pH, the dihedral angle of the diol, the
buffer and its ionic strength, the solvent, and the stability

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DOI: 10.1002/9780470661345.smc014

Concepts

OH
B
OH

O
B

1
Protic
solvent

Protic
solvent

HO
+

2H2O

HO

+ 2H2O

Protic
solvent

Protic
solvent
H

Kapp

OH
B OH
OH

O
B
O
OH

Scheme 1

Overall binding equilibrium of phenylboronic acid with a diol.

of the boronic acid. These issues have been comprehensively discussed in a review by Wang and coworkers, and
is not dealt with in detail here.3 Mentioned below are a few
representative examples that show the use of the boronic
aciddiol interactions.
Since the publication of the seminal papers on boronic
acid-based carbohydrate recognition by Czarnik5 and
Shinkai6 in the early 1990s, the field has undergone very
significant growth.4 Earlier efforts were mostly focused
on recognition of simple sugars such as mono- and disaccharides.1, 7 Later on, there were extensive efforts in
the development of boronic acids capable of fluorescent
property changes upon binding.4 These areas have been
covered in depth in recent reviews3, 4, 8 and are not discussed. Recent efforts have moved to the recognition of
biologically important carbohydrate biomarkers and building lectin mimics (boronolectins) using DNA, peptides,
and proteins scaffolds. Along these lines, there have been
efforts in the development of bisboronic acids for the
recognition of cell-surface carbohydrate biomarkers in cancer,8 peptide-based lectin mimics for the recognition of
the T-antigen,9 boronic acid-modified proteins,10, 11 and
boronic acid-modified DNA for various applications.12, 13
Below, selected, recent examples are described to illustrate
the utility of the boronic acid moiety for recognition of biologically important carbohydrates and in other applications.
Glycoprotein is defined as proteins that contain oligosaccharides covalently attached to polypeptide side chains. The
process is also called glycosylation. It is well known that
glycosylation plays a critical role in governing the function
and activity of many proteins.14, 15 Although the mechanism of glycosylation is well understood, the rapid detection
and differentiation of different glycosylation patterns in the
same protein is not a trivial issue. Tools commonly used
include antibodies16 and aptamers.1719 However, these
methods only focus on the detection of the intact protein

without the ability to specifically probe differences in glycosylation. To address this issue, the Wang group is working on incorporating boronic acid-modified thymidine into
DNA for aptamer selection work for glycoproteins. The
central hypothesis is that, due to the intrinsic binding affinity of boronic acids with carbohydrates, incorporation of
a boronic acid into the DNA aptamer would gravitate the
selection process toward the glycosylation site. The general aptamer selection method and idea were developed
about 20 years ago by the labs of Szostak,17 Joyce,19 and
Gold.18 Specifically, in the modified approach used in the
Wang lab, 8-quinolynylboronic acid-modified thymidine5 -triphosphate (QB-TTP, Scheme 2) was first synthesized
and effectively incorporated into DNA through polymerasemediated DNA synthesis.12 Recently, a second analog of
QB-TTP (NB-TTP, Scheme 2) was also reported by the
Wang group.13 Compared to the first-generation of boronic
acid-modified TTP (QB-TTP), NB-TTP has its own unique
properties. It changes fluorescent properties upon sugar
binding (Ka = 73 M1 with D-fructose) and the fluorescent
properties are retained after DNA incorporation (1.5-fold
change of fluorescence intensity upon addition of 100 mM
D-fructose). With such fluorescent properties, NB-TTP can
be used in applications such as fluorescent DNA aptamer
selection, DNA labeling work, as well as genomic DNA
incorporation.
Recently, there also have been efforts in making peptidebased boronic acid lectin mimics (PBLs) for glycoproteins. For example, Lavigne and coworkers selected their
PBLs for the detection of glycoproteins according to
response patterns by arrays of unidentified peptideboronic
acids libraries20 ; the Duggan group studied 4-borono-Lphenylalanine-based PBLs and their affinity for alizarin21 ;
and the Anslyn group reported a chemosensor array of PBLs
for saccharides and their derivatives. Such arrays were used
for the identification of sucralose from a beverage sample.22

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Reversible covalent bond toolbox

O
N3

N
H
O

or

N3
O

HO B OH
QB

NB
HN

H
N

O
NH

O
HO

OH OH OH
O O O
P
P
P
O O O

N N
N

R=

O
QB-TTP

O
NH

O
OH OH OH
O O O
P
P
P
O
O O

HO B OH

OH
H
N

HO

OH
B
OH

N
H

O
O

O
OH

R=
NB-TTP
HN

Scheme 2

N N
N

OH
B
OH

Synthesis of BQ-TTP and NB-TTP.

As a specific example, a recent study by the Hall lab

is used for in-depth discussions. As a prelude, in 2006,
the Hall lab first reported ortho-hydroxymethyl phenylboronic acid (5, Figure 1), which has a weak but encouraging binding with a model of hexopyranosides (methyl
-D-glucopyranoside).23, 24 This is especially important,
since carbohydrates found in glycoproteins, glycolipids, and
lipopolysaccharides are almost always six-membered ring
sugars and linear diols, while most boronic acids normally
show a preference for furanose sugars in binding. Compound 5 binds methyl -D-glucopyranoside with a Ka of
22 M1 in neutral aqueous media by complexing hexopyranosides primarily using their 4,6-diol. Having this compound on hand, the Hall group recently developed a library
of PBL by incorporating 5 as hexopyranoside-binding
agents.9 After competitive enzyme-linked immunosorbent
assay (ELISA) screening of the library, compound 6 was
selected as the best receptor for the ThomsenFriedenreich
(TF) antigen (Kd = 0.9 mM in neutral water), which is
implicated in cancer. Although further studies such as
exploiting the possibility of selectively labeling TF-specific
tumor cells is needed to demonstrate full utility, this is an

O
B

OH

O
B

HO

O
O
H2N

N
H

HN
H
N
O

HN
O
N
H

CH3O

H
N
O

N
H

B O

Building block:

Figure 1

HO

Structures of compound 5 and PBL 6.

excellent example developing PBLs for carbohydrate-based

biomarkers.
In addition to PBLs mentioned above, the Schultz
lab also reported boronic acid-modified proteins. Specifically, the Schultz lab successfully incorporated a boronic
acid-modified phenylalanine (p-boronophenylalanine, 7,
Figure 2) into protein for the selection of lectin mimics with

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Concepts

3.1

Interactions with amino groupsreversible

Schiff base formation

OH
HO

Figure 2

B
OH

NH2

Reversible covalent interactions between a carbonyl group

and an amino group result in the formation of a Schiff
base (imine) or an aminol. There have been a number
of research reports on taking advantage of such reactions
for sensing.25, 2938, 40, 48 For example, a coumarin aldehydebased sensor for amines was developed by the Glass group
(8, Scheme 3).35 The coumarin aldehyde 8 displayed a large
(34 nm) red shift in absorption and a strong fluorescence
increase when exposed to glycine. It is believed that the
hydrogen bond between the iminium ion and the carbonyl
group of the coumarin effectively modulates the coumarin
fluorescence, thus inducing the change in its spectrometric
property.
A series of similar sensors were also developed by
the Glass group using dimers of a quinolone aldehyde
chromophore for the recognition of diamines.31 Good

Structure of p-boronophenylalanine 7.

enhanced affinities for carbohydrates.10, 11 This strategy can

also be extended to other uses such as purification of native
protein sequences by a one-step scarless affinity procedure
as well as in vivo labeling of boronate-containing proteins.

ELECTRON-DEFICIENT CARBONYL
GROUPS AND THEIR INTERACTIONS
WITH HYDROXYL GROUPS AND
AMINO GROUP

The carbonyl group has always been an important functional group in molecular recognition. It readily reacts with
various nucleophiles, among which some are reversible.
Along this line, there have been efforts in the development of chemosensors based on reversible covalent
interactions between electron-deficient carbonyl groups
and different nucleophiles, such as hydroxyl groups,2527
amino groups,25, 2840 bisulfite,41 sulfide,42 hydrozine,43 and
cyanides.4447

R
O

H3 N

Bu

Bu H
O

O
N

S
S

Bn
N O

H
N

H2N

O
O

OH

S
S

H
N

N O
Bn
11

10

or

or

Scheme 4

or

or

or

O
N
H

N O
Bn

X=

or
f

NH2

Scheme 3 Reversible interaction between coumarin aldehyde 8

and amino acid.

Bn
N O

N
H

Amino acid sensors based on imine formation.

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Reversible covalent bond toolbox

in the detection of amines with a distinct color change

upon binding (50 nm blue shift). Thus, a series of amine
sensors have been developed by Mohr and coworkers
(Figure 3, ETHT 40014014).25, 29, 30 Recently, a solgelbased optical sensor for the detection of amines in aqueous solution has been developed using ETHT 4001 as the
indicator dye.51 The sensor system shows good reversibility and fast response to the exposure of amines (e.g.,
propylamine), and has potential applications in the food
industry.
The trifluoroacetylamine interaction has also been used
in creating artificial receptors using the molecular imprinting method. For example, 1,4-diaminobutane was imprinted
into polymer prepared using a dendrimer dye as the
monomeric unit, which included trifluoroacetyl azobenzene as the chromogenic reporter for amine.34, 48 After
the dendrimer was cross-linked using the Grubbs catalyst,
template 1,4-diaminobutane was removed, which presumably left behind a complementary cavity. The imprinted
polymer selectively binds to the aliphatic diamine with
a C3 or C4 carbon chain between the two amino groups
(Scheme 7) with an apparent association constant (Kapp =
2.7 104 M1 ), which is 200-fold higher than that for
butylamine.

binding constants and large fluorescence changes (30 nm

red shift) were observed. Different linker lengths were
utilized to optimize selectivity and binding with various
diamines (10ag, Scheme 4).
Among these series of sensors, there is one that takes
advantage of an electron-deficient carbonyl group and a
phenylboronic acid moiety in a bifunctional fluorescent
sensor (Scheme 5).32 The boronic acid-containing coumarin
aldehyde 12 binds selectively with catecholamines such as
dopamine and norepinephrine as a result of Schiff base
formation, the intrinsic affinity of the boronic acid toward
the catechol diol group, and the specifically designed threedimensional arrangement. A similar design from the Yoon
group describes an anthracene bearing a phenylboronic acid
moiety and an aldehyde carbonyl group for sensing of
dopamine.49
Similar types of interactions can also be used in the
detection of aldehydes. A fluorescent chemosensor reported
by the Mohr group exhibits a turn-on fluorescence
property upon exposure to an aldehyde (Scheme 6).36
The reversible trifluoroacetophenoneamine interaction50
has been extensively studied. On the basis of this type of
interactions, fluorescent compounds such as trifluoroacetylstilbene27, 28, 40 and trifluoroacetylazobenzene29 have used

NH

OH

B(OH)2
O
N

12

HO

NH2

N
H
13

HO
O
N

O
N

B(OH)2

B O
OH

NH
14

Scheme 5

15

Catecholamine sensors utilizing both aminealdehyde and boronic aciddiol interactions.

O
C6H13
C6H13

N NH2

R1

16 (Non-fluorescent)

Scheme 6

R2

O R2

N N

C6H13
C6H13

R1

17 (Fluorescent)

Reversible interaction of compound 16 with aldehyde.

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Concepts

R1
N
R2

N
N

CF3

18 R1 = R2 = C8H17, ETHT 4001

19 R1 = C11H22OH, R2 = C2H5
OC11H22
, ETHT 4012

20 R1 =

R3
N
R4

CF3

22 R3 = R4 = C8H17, ETHT 4004

23 R1 = R2 = C4H9, ETHT 4003
OC11H22

24 R1 = R2 =

, ETHT 4014

O
R2 = C2H5
21 R1 =

OC11H22
Polymer

R2 = C2H5

Figure 3

Structures of compounds 1824.

N
N

F3C

O
O

compounds containing hydroxyl groups, there have been

only a limited number of studies of such sensors based
on carbonyl groups. However, some sensors developed for
amines have similar responses to alcohols, thus posing
an interference problem.2527 The same strategy has been
extended to chemosensors for other nucleophiles, such as
sulfur nucleophiles42, 52 and cyanide.4547, 53

3.3

Interactions with sulfur nucleophiles

Diamine
O
CF3

N
N

Scheme 7 A cartoon description 1,4-diaminobutane binding to

imprinted polymer.

3.2

Interactions with hydroxyl groups

Similar to amino groups, hydroxyl groups can also engage

in reversible covalent interactions with electron-deficient
carboxyl groups. Therefore, it is conceivable that carbonylbearing fluorophores could also be used in the development of sensors for molecules containing hydroxyl groups.
However, due to the wide use of boronic acids in sensing

Detection of sulfide in an aqueous solution is very useful in the food industry. The MartinezManez lab has
developed a highly selective colorimetric sensor for sulfide anion by taking advantage of the pyrylium cycle in a
2,4,6-triphenylpyrylium analog.42 This reaction is shown
in Scheme 8. The pyrylium ring is formed when the pH of
the solution is adjusted to acidic conditions. After the addition of sulfide at only 5 105 M, the color of the solution
changes dramatically from magenta to blue, which is easily detectable by the naked eye. This is explained by the
formation of the thiopyrylium ring (27).
The reversible covalent interactions between an aldehyde
and hydrogen sulfite are useful for detection applications
in the wine industry, where hydrogen sulfite is used to
stabilize wine during/after fermentation. As an example,
the colorimetric sensor for hydrogen sulfite developed by
the Mohr group52 possesses an alkylamino azobenzene
backbone, which is similar to the amine sensors developed
by the same group. Instead of the trifluoroacetyl group,
formyl group is used to bind with the bisulfite anion. A
36-nm red shift (from 524 to 488 nm) was observed when
the compound binds to bisulfite anion, leading to a color
change from pink to orange (Scheme 9).

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Reversible covalent bond toolbox

OH

SH, H+

OH

OH

25

Scheme 8

H+

26 (Magenta)

27 (Blue)

Sulfide anion sensor based on the pyrylium cycle.

O2N
C8H17
C8H17

N
N

O2 N

O
H

HSO3 C8H17

3.4

OH
SO3H
H

N
N

C8H17
28 (Pink)

Scheme 9

29 (Orange)

Bisulfite sensing using an azobenzene derivative.

Interactions with cyanides

The use of trifluoroacetophenoneanion interactions is

not limited to the sensing of amines. Chemosensors for
cyanides have also been developed on the basis of the trifluoroacetophenone structure.4447 For example, the Ahn
group has reported four different types of sensors for
cyanides (Scheme 10). Though these four sensors take
advantage of different types of chromophores and fluorophores, they all include the o-(carboxamido) trifluoroacetophenone moiety in their structures. In the first
example,45 oligothiophene was used as the fluorophore.
Upon exposure to cyanide or acetate, the nonfluorescent
compound 30 became fluorescent (31), with a large fluorescence enhancement factor (FEF) of 120. Such findings are very valuable because, in most others cases,
binding of an anion (e.g., acetate) often leads to fluorescence quenching rather than enhancement. The enhancement in this case is due to the elimination of the quenching processes on the fluorophore by the addition of the
anionic guest. In the second example in Scheme 10,47 the
DNPA (2-(2,4-dinitrophenylazo)-1-naphthol-3,6-disulfonic
acid) sensor showed a significant intensity change in
absorption in the 433533 nm range upon binding to
cyanide, leading to an orange-red color in the aqueous
solution (H2 OCH3 OH). In contrast, no response was
observed toward other reference anions, such as fluoride, acetate, phosphate, sulfide, thiocyanide, chloride, and
bromide. This is explained by the fact that anions such
as F , AcO , and H2 PO4 2 are stabilized by solvation
effect, whereas CN , which is a poor H-bond acceptor,
shows strong nucleophilicity and attacks the carbonyl group
more readily, leading to the consequent spectrometric
property changes. The high selectivity toward cyanide and

CN

H N
O

H N
O
F3C CN

F3C
30 (Non-fluorescent)

31 (Fluorescent)

NO2

NO2
H
N

O2N

CF3

H
N

CN

O2N

CF3
CN

32 (Colorless)

33 (Orange-red)

N
CN

H
O S
O
O N

O S H
O
O N
CF3

CF3
CN

34

35 (Increased fluorescence
intensity)
O

Et
N

CN

H
O

CF3
36

Scheme 10

O
CN

Et
N

CN

H
O

NC

CF3
37 (Decreased fluorescence
intensity)

Trifluoroacetophenone-based sensors for cyanide.

the convenient detection by naked eye in aqueous media

make this agent very useful. The third44 and fourth46

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Concepts

(NO3)4
Zn2+

Zn2+
N

Zn2+

O
P
O
O

2+

O O
P
O
O

Zn2+

Phosphorylated
peptide

Phosphorylated
peptide

Sensor for phosphorylated site of peptide based on DPAZn.

examples in Scheme 10 are similar in terms of the binding

mechanism. An interesting aspect in these examples is that
the fluorophore substitution on the carboxamido group and
the phenyl ring can both result in spectrometric property
modulation, indicating that the anion-H-bond stabilization
process can affect the electron density distribution on both
parts. This versatility in modification and modulation offers
the possibility of developing new sensors with diverse
structural and spectroscopic properties.

Zn

N
38

Figure 4

Zn2+
O

METALANION/LIGAND
INTERACTIONS

The selective recognition of anions is an important

research field. This is because most of the important
biomolecules, such as nucleotides, peptides, phospholipids,
and some carbohydrates, are anionic. However, anion
recognition in aqueous solution has always been challenging because the dominant solvation effect keeps all
anions strongly hydrated. The comparatively large and
complicated structure of anions also makes selective and
efficient sensing difficult. Although reversible interactions
between electron-deficient carbonyl groups and anions
have been utilized in sensor design, much more work
is needed in order to achieve high sensitivity and selectivity. Nature has employed metalanion/ligand interactions in molecular recognition involving proteins/enzymes,
nucleic acids, and phospholipids. Such interactions have
also been used by scientists in the development of
binders for anions with impressive results.5464 These
sensors are composed of a fluorescent or nonfluorescent multivalent ligand, which coordinates with one or
two metal cations, such as Zn2+ , Cu2+ , and Cd2+ , to
form a ligandmetal complex. Besides, fluorescent lanthanides, such as Tb and Eu have also been used in
some sensors, especially those made by the Gunnlaugsson
group.5456, 61 In terms of detection, there are two major
types of fluorometric or colorimetric sensing mechanisms,

namely, indicator displacement mechanism and fluorescence quenching mechanism.

Many of the sensors based on dimetallic coordination
complexes take advantage of the dipicolylamine (DPA)
ligand, which could form stable complexes with various
metal cations. The first example is from the Hamachi group
(38, Figure 4).64 This fluorescent chemosensor selectively
binds to phosphorylated peptides with both high affinity
(107 M1 ), giving significant fluorescence increase (45fold). In contrast, it does not respond to nonphosphorylated
peptides. Because of the enormous importance of protein
posttranslational phosphorylation, this type of phosphate
sensors has become very useful in the detection and/or
understanding of protein phosphorylations.
Along a similar line, the Hong group designed a series of
ZnDPA-based sensors for pyrophosphate (Figure 5).65, 66
The two sensors in Figure 5 are both based on the dimetallic complex, and the phenolate oxygen is utilized to bind
with Zn and establish a connection between the pyrophosphate binding and the electron density in the fluorophore.
Compound 39 is a colorimetric sensor that shows a red shift
of about 50 nm (color from yellow to red) upon addition of

NO2

Zn2+

N
Zn2+

N
N

Figure 5
phate.

OO N
P
O
O P O
O
39
O

N
N

Zn2+

N
Zn2+
N

N
N

OO N
P
O
O P O
O
O

40

Colorimetric and fluorometric sensors for pyrophos-

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Reversible covalent bond toolbox

F3C

R
N

41

Figure 6

(ClO4)2
N

43 (Weakly fluorescent)

Scheme 11

inorganic pyrophosphate (PPi). This sensor also displays

high selectivity for PPi compared to other anions including
Pi. Compound 40 is a fluorometric sensor that shows a 9.5fold fluorescence enhancement upon addition of PPi and
good selectivity for PPi compared to ATP, ADP, AMP,
and inorganic phosphate (Pi). It is believed that the binding
of PPi to the Zn cation weakens the bond between Zn and
the phenolate oxygen, which then becomes more negatively
charged and induces colorimetric changes in the azobenzene
fluorophore. More dimetallic chemosensors are reviewed in
detail by Smith.67
The indicator displacement principle has been utilized
in many sensor designs. The indicator(s) can be incorporated into the sensor itself, or they may be independent
fluorescent molecules. Two different sensors have been
developed by the Kikuchi and the Mohr labs, respectively. The sensor developed by the Kikuchi group63 is a
Cd(II) complex, which binds to AMP much more strongly
than cAMP, and gives a 39 nm red shift in fluorescence
upon binding (Figure 6). Because of this interesting character, this sensor might be useful in monitoring the activity of phosphodiesterase, which converts cAMP to AMP.
The sensor recently reported by the Mohr group68 has
a Zn(II) core coordinated by a DPA unit, which is further linked to a naphthalimide fluorophore. When the ATP
anion comes to bind with the Zn to displace the imine,
fluorescence is greatly enhanced as a result of a decrease
in quenching resulting from photo-induced electron transfer (PET).

Target
protein

Ni

NO2
44 (Strongly fluorescent)

NBD-based sensor for Zn(II).

Another very interesting example came from the Wang

group, which developed an NBD (7-nitrobenzo-2-oxa-1,3diazol) based fluorescent probe (43, Scheme 11) for zinc
ion.69 This probe is water-soluble and very sensitive to
Zn(II) (Kd = 4.6 M) with 25-fold maximum fluorescence
intensity change in phosphate based saline (PBS) buffer
(pH = 7.3). Besides, 43 also showed very good selectivity
against other represented metal ions in living cells, such
as Na+ , K+ , Mg2+ , Ca2+ , Mn2+ , Fe2+ , Fe3+ , Cd2+ , Ni2+ ,
Co2+ , Hg2+ , and Cu2+ .
Reversible metalanion interactions have also been used
in affinity purification of proteins. For example, immobilized Ni2+ absorption chromatography has been used to
purify histidine-tagged proteins.70 In this approach, the protein of interest that needs to be purified is modified using a
His-6 label or His-tag, that is, a polyhistidine residue tagged
on the amino or carboxyl terminus, upon expression. Such
proteins can be easily separated from the cell lysate mixture through reversible His-6-Ni interactions. The protein
of interest can be recovered in a pure state (>95%) by
washing the resin with high concentrations (50100 mM)
of imidazole (Figure 7). This is among the most popular
and successful applications of reversible metalanion interactions in molecular recognition.
From the above examples, one can see that metalanion
interactions have proven to be very useful in the development of sensors for anionic species, especially biomolecules
containing the phosphate group. However, sensors with
higher selectivity and sensitivity for similar or other biologically important targets are still very much needed.

Binding
washing

Figure 7

NO2

Indicator displacement sensors for AMP and ATP.

NH
O

Zn2+N

N
2+
Zn N
N

42

Target
protein

Zn2+

HN

NH

NH N
Cd
NH N
H

OH

R=
HN

Ni

Target
protein
Imidazole
elution

Affinity purification of protein based on NiHis-tag interaction.

Supramolecular Chemistry: From Molecules to Nanomaterials, Online 2012 John Wiley & Sons, Ltd.
This article is 2012 John Wiley & Sons, Ltd.
This article was published in the Supramolecular Chemistry: From Molecules to Nanomaterials in 2012 by John Wiley & Sons, Ltd.
DOI: 10.1002/9780470661345.smc014

10

Concepts

O
R'SH

R'S

N
R

O
45

O
46
O

R=

or

Bu
N

and other nucleophiles/Lewis bases, (ii) interactions of

electron-deficient carbonyl groups with hydroxyl groups
and amino group or other nucleophiles, and (iii) metal
anion/ligand interactions. The extraordinary strength and
their readily reversible nature put these covalent interactions
in a very unique position for the construction of receptors
of high affinity and specificity.

or

ACKNOWLEDGMENTS
NH

Scheme 12

Reactions of sensor 45 with a thiol.

Work conducted in the authors lab was supported by the

National Institutes of Health (GM086925 and GM084933)
and Georgia State University Molecular Basis of Disease
(MBD) program (YFC).

CHEMOSENSORS BASED ON THE

MICHAEL ADDITION REACTION
REFERENCES

Michael addition is one chemical reaction that has been

used for the development of chemosensors. In reality, in
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reagents than sensors because Michael additionbased
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the fluorophore and the electron-deficient alkene. However,
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yield changing from 0.19 to 0.540.61 after addition of
a thiol. It is well suited for optical sensing since both
absorbance and emission wavelengths are in the visible
spectral range (ex : 443 nm, em : 525 nm).

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Supramolecular Chemistry: From Molecules to Nanomaterials, Online 2012 John Wiley & Sons, Ltd.
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DOI: 10.1002/9780470661345.smc014