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10-11-2016
Recent approaches in designing bioadhesive materials inspired by Recent approaches in designing bioadhesive materials inspired by
mussel adhesive protein mussel adhesive protein
Pegah Kord Forooshani
Michigan Technological University
Bruce P. Lee
Michigan Technological University
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Recommended Citation Recommended Citation
Forooshani, P. K., & Lee, B. P. (2016). Recent approaches in designing bioadhesive materials inspired by
mussel adhesive protein.
Journal of Polymer Science, Part A: Polymer Chemistry
. http://dx.doi.org/
10.1002/pola.28368
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https://digitalcommons.mtu.edu/biomedical-fp
Part of the Biomaterials Commons, Polymer Chemistry Commons, and the Polymer Science Commons
Recent Approaches in Designing Bioadhesive Materials Inspired
by Mussel Adhesive Protein
Pegah Kord Forooshani, Bruce P. Lee
Department of Biomedical Engineering, Michigan Technological University, Houghton, Michigan 49931
Correspondence to: B. P. Lee (E-mail: bplee@mtu.edu)
Received 1 August 2016; accepted 3 September 2016; published online 00 Month 2016
DOI: 10.1002/pola.28368
ABSTRACT: Marine mussels secret protein-based adhesives,
which enable them to anchor to various surfaces in a saline,
intertidal zone. Mussel foot proteins (Mfps) contain a large
abundance of a unique, catecholic amino acid, Dopa, in their
protein sequences. Catechol offers robust and durable adhe-
sion to various substrate surfaces and contributes to the curing
of the adhesive plaques. In this article, we review the unique
features and the key functionalities of Mfps, catechol chemis-
try, and strategies for preparing catechol-functionalized poly-
mers. Specifically, we reviewed recent findings on the
contributions of various features of Mfps on interfacial binding,
which include coacervate formation, surface drying properties,
control of the oxidation state of catechol, among other fea-
tures. We also summarized recent developments in designing
advanced biomimetic materials including coacervate-forming
adhesives, mechanically improved nano- and micro-composite
adhesive hydrogels, as well as smart and self-healing materi-
als. Finally, we review the applications of catechol-
functionalized materials for the use as biomedical adhesives,
therapeutic applications, and antifouling coatings.
V
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2016 The
Authors. Journal of Polymer Science Part A: Polymer Chemis-
try Published by Wiley Periodicals, Inc. J. Polym. Sci., Part A:
Polym. Chem. 2016, 00, 000–000
KEYWORDS: adhesives; adhesive polymers; biomaterials;
biomimetic design; biopolymers; Dopa; mussel foot proteins;
wet adhesion
INTRODUCTION
Marine mussels have mastered the ability to
anchor to foreign surfaces in seawater through the use of adhe-
sive proteins.
1
These mussel foot proteins (Mfps) are known to
cure rapidly to form adhesive plaques with high interfacial
binding strength, durability, and toughness. 3,4-Dihydroxyphe-
nylalanine (Dopa), which is modified from tyrosine through
post-transitional hydroxylation, is one of the main constituents
in Mfps.
2–4
The catechol side chain of Dopa has the ability to
form various types of chemical interactions and crosslinking,
which imparts Mfps with the ability to solidify in situ and bind
tightly to various types of surface substrates. To harvest the
remarkable wet adhesive properties of these adhesive proteins,
many efforts have been made to develop new adhesive materi-
als inspired by the designs of Mfps.
Natural Mfps have been extracted and analyzed from differ-
ent species of mussels in the aim of creating strong adhesive
materials.
5,6
However, several thousand mussel specimens
are required for extracting one gram of adhesive proteins,
making the direct use of these adhesives for commercial
applications highly challenging.
7
This highlights the need for
developing synthetic mussel-inspired adhesive polymers
with strong water-resistant adhesive properties.
The adhesive mechanism of marine mussels and the key fea-
tures of Mfps that affect adhesive properties have been
extensively studied during past decades, which provide guid-
ance for developing new synthetic biomimetic adhesives.
8–10
Here, we provide an updated review on the design of adhe-
sive materials inspired by Mfps. We first describe the unique
features of adhesive plaque proteins and their key function-
alities as well as strategies for preparing biomimetic adhe-
sive polymers. We also summarize the recent developments
in designing advanced mussel-inspired materials including
coacervated Dopa-functionalized adhesives, mechanically
improved nano- and micro-composite adhesive hydrogels,
hydrogel actuators, self-healing hydrogels, and smart adhe-
sives. Finally, the applications of these adhesive materials as
biomedical adhesives, drug carriers for therapeutic uses, and
antifouling coatings are reviewed.
CHEMISTRY OF ADHESION: MUSSEL ADHESIVE PROTEINS
Mussel adhesives proteins enable marine mussels to attach
strongly to various surfaces in their turbulent, wet and saline
habitats.
1
These proteins are secreted in a liquid form, which
then solidify to form a byssal thread and an adhesive plaque
V
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2016 The Authors. Journal of Polymer Science Part A: Polymer Chemistry Published by Wiley Periodicals, Inc.
This is an open access article under the terms of the Creative Commons Attribution-NonCommercial-NoDerivs License, which
permits use and distribution in any medium, provided the original work is properly cited, the use is non-commercial and no
modifications or adaptations are made.
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complex (Fig. 1). The byssal threads are engineered to with-
stand elevated mechanical loads applied by waves and cur-
rents. A byssal thread connects a mussel to the adhesive
plaque that is anchored to a foreign surface.
2,11
The average
force needed to dislodge a California mussel, Mytilus califor-
nianus, is estimated to be 250–300 N/mussel, with an average
detachment force of 5–6 N/thread.
12,13
This remarkable sur-
face anchoring capacity provides insights in designing synthet-
ic polymers for interfacial applications. In this section, we
review the compositions and chemistries of various plaque
proteins with a specific focus on those found at the plaque-
surface interface that contribute to strong interfacial binding.
Mussel Foot Proteins
At least six Mfps (Mfp-1 through Mfp-6) have been identified
from the adhesive plaques of several species of mussel (i.e.,
Mytilus edulis, Mytilus galloprovincialis, M. californianus,
etc.).
14
These proteins are characterized by a basic isoelectric
point (pI) due to a high content of cationic amino acids. The
pI is described as the pH in which the net charge of the pro-
tein is zero.
15
Most importantly, these plaque proteins con-
tain various amounts of the unique amino acid, Dopa.
3,4
The
catechol side chain of Dopa offers robust and durable adhe-
sion to various substrate surfaces and contributes to the cur-
ing of the adhesive plaques.
4
Mfps extracted from different
Mytilus species exhibited a high level of sequence homology
and a similar distribution within the adhesive plaque
(Fig. 1).
12,13
These findings suggest tha t each type of Mfp
has a unique function and contributes differently to the
interfacial properties of the adhesive plaque.
Mfp-1 is a high molecular weight (e.g., 108 kDa in M. edulis)
and basic protein with very little secondary structures.
16
It
is located in the cuticle of the byssus threads and the adhe-
sive plaques and acts mainly as a protective varnish layer.
17
Mfp-2 is a smaller protein (e.g., 42–47 kDa in M. edulis) with
highly repetitive motifs and is the most abundant protein
found within the plaque (25 wt %).
18
Mfp-2 contains a rel-
atively high content of cysteine residues (6 mol %) in the
form of disulfide bonds, and it is believed that Mfp-2 pro-
vides structural integrity to the adhesive plaques.
18
Mfp-4
consists of a histidine- (His-) rich decapeptide tandemly
repeated more than 36 times, which binds exceptionally well
to transition metal ions such as copper.
19
Mfp-4 is strategi-
cally located between the adhesive plaque and the distal por-
tion of the byssal thread, effectively linking plaque proteins
(e.g., Mfp-2) with those found within the byssal thread (e.g.,
preCOL).
9,19
PreCols are collagenous proteins with high Dopa
and His contents and these proteins are mainly distributed
throughout the byssal threads.
20,21
Specifically, Mpf-4 is
believed to interact with the His-rich domain of preCOLs
through metal ion coupling.
9,19,22
Mfp-3, 5, and 6 are predominantly found at the plaque–sub-
strate interface, contributing to strong, wet adhesion. Mfp-3
is the smallest adhesive protein among plaque proteins (e.g.,
molecular weight of 5–7 kDa in M. edulis and M. california-
nus).
22–24
It is the most polymorphic adhesive protein with
no known repeating sequences.
24
It is reported to have over
30–35 different variants, which can be further subdivided
into two separate groups known as Mfp-3 fast and slow
(Mfp-3f and Mfp-3s, respectively).
24,25
Based on the sequen-
ces reported for M. californianus (Table 1), both Mfp-3f and
Mfp-3s are rich in glycine (25–29 mol %) and asparagine
(10–18 mol %).
24
Additionally, Mfp-3f exhibits elevated con-
tents of post-translationally modified Dopa (> 20 mol %)
and 4-hydroxyarginine (1 mol %), and positively charged
residues (26 mol %) [Fig. 1(A,E)], which renders Mfp-3f
highly hydrophilic.
In contrast, Mfp-3s exhibits a lower conversion of tyrosine to
Dopa residues (5–10 mol %) and contains a lower charge
density (9 and 3 mol % positively and negatively charged
residues, respectively) when compared to Mfp-3f, resulting
in a polar but hydrophobic protein [Fig. 1(B)]. However, its
Dopa content can approach 28 mol % in some variants,
which may be crucial for adhesion to metal and mineral sur-
faces.
24
This level of Dopa content is only exceeded by
another plaque protein, Mfp-5.
26
In Mfp-3s, Dopa is
Pegah Kord Forooshani received her bachelor’s degree in chemical engineering from Uni-
versity of Tehran, Iran, and her master’s degree in biomedical engineering from University
of Malaya, Malaysia. During her master studies, she worked on the synthesis and charac-
terization of citric acid-based polyester elastomers for tissue engineering applications. Cur-
rently, she is perusing a PhD degree in biomedical engineering at Michigan Technological
University, USA, under the supervision of Professor Bruce P. Lee. She focuses on tuning
the redox chemistry of catechol for promoting wound healing.
Prof. Bruce P. Lee received a PhD degree in biomedical engineering from Northwestern
University. Prior to his current appointment at Michigan Technological University, he co-
founded a startup company, Nerites Corporation, which aimed at commercializing adhe-
sives and coating inspired by mussel adhesive proteins. His current research interests lie
in utilizing the interfacial chemistries of these adhesive proteins in designing bioadhesives
and smart materials. He was awarded the 2016 Young Investigator Award from the Office
of Naval Research.
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protected from oxidation even in footprints left behind from
the removed plaques. This is surprising given the susceptibil-
ity of the Dopa to autoxidation at the pH of seawater (pH
7.5–8.4).
27
We reviewed the possible reduction–oxidation
(redox) chemistry of plaque proteins in the subsequent sec-
tion entitled “Effect of Oxidation State of Catechol.”
Mfp-5 has a molecular mass of 8.9 kDa and is the least poly-
morphic plaque proteins, consisting of one protein sequence
with two closely related variants.
14,26
Mfp-5 contains the
highest amount of the adhesive Dopa (30 mol %) residues
amongst all the plague proteins [Fig. 1(C)].
14
Mfp-5 is also
characterized by its hydrophilicity and a basic pI due to an
FIGURE 1 Schematic representation of a byssal thread and adhesive plaque with the approximate distribution of known Mfps. Pri-
mary sequences of Mfp-3f (A), Mfp-3s (B), Mfp-5 (C), and Mfp-6 (D). Acidic, basic, Dopa, and aromatic residues are shaded blue,
red, dark purple, and light purple, respectively. Post-translational modification of tyrosine to Dopa and the formation of disulfide
cysteine could occur anywhere within the peptide sequences. Pie charts illustrating the distribution of key functionalities found in
selected Mfps (E).
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elevated content of cationic amino acids (27.7 mol %). Addi-
tionally, Mfp-5 also contains variable amounts of post-
translationally modified phosphoserine (4.8 mol %), known
for its ability to bind to calcareous mineral materials (e.g.,
statherin and osteopontin).
28,29
The presence of elevated
Dopa content and phosphoserine suggest that Mfp-5 plays
an important role in interfacial binding.
Unlike aforementioned interfacial plaque proteins, Mfp-6
contains a much lower amount of Dopa (3 mol %) but a
higher levels of tyrosine (20 mol %) [Fig. 1(D)].
14
Although
the total phenolic residues content in Mfp-6 is similar to
those found in Mfp-3 and 5, the tyrosine residues in Mfp-6
are not efficiently converted to Dopa. Additionally, Mfp-6 has
the highest contents of charged residues (23 and 16 mol %
anionic and cationic amino acids, respectively). Another
unique feature of Mfp-6 is the presence of cysteine (11 mol
%) with a small portion of these residues present in the
form of disulfide bonds (2 mol %). The high level of thiol
gives Mfp-6 the ability to effectively control the redox chem-
istry of Dopa residues present in other plaque proteins,
14,30
which is further reviewed in a later section.
Catechol Chemistry
One of the unique features of Mfps is the abundance of the
catecholic amino acid, Dopa, in their protein sequences. The
presence of catechol is believed to fulfill the dual role of inter-
facial binding and the solidification of the adhesive proteins.
31
Catechol is capable of diverse chemistries, which enables it to
bind to both organic and inorganic surfaces through the
formation of reversible non-covalent or irreversible covalent
interactions (Fig. 2). These chemical reactions are also critical
to designing in situ curablematerials.Inthissection,wesum-
marize various catechol side chain chemical interactions.
Non-Covalent Dopa Interactions
The dihydroxy functionality of catechol enables it to form
strong hydrogen bonds [H-bonds, Fig. 2(A)], which promotes
its absorption to mucosal tissues
32,33
and hydroxyapatite
surfaces.
34,35
The benzene ring of catechol is capable of
interacting with other aromatic rings through p–p electron
interaction [Fig. 2(B)],
1,11
which improves the cohesive prop-
erties of catechol-containing polymers and enables them to
attach to surfaces rich in aromatic compounds (e.g., polysty-
rene)
36
and gold substrates.
37
The aromatic ring also forms
cation–p interaction with positively charged ions, which is
one of the strongest non-covalent interactions in water [Fig.
2(C)].
38–40
Cation–p interaction enhances absorption of cate-
chol to charged surfaces
41
and contributes to the cohesive
properties of materials rich in both aromatic and cationic
functional groups.
42
Since catechols are easily oxidized to its
poorly adhesive quinone form in an oxygen rich and basic
environment, cation–p interaction complements the under-
water adhesive properties of catechol.
42–45
Catechol chelates metal ions to form strong, reversible com-
plexes with various metal ions, including Cu
21
,Zn
21
,Mn
21
,
Fe
31
,V
31
,Ti
31
,andTi
41
[Fig. 2(D)].
46–49
The log stability con-
stants of these complexes are significantly higher when com-
pared to those of polymeric acid- or amino acid-based ligands
TABLE 1 Amino Acid Composition of Mussel Foot Proteins Isolated From the Plaque Represented in the Number of Residue Per
100 Residues
Amino acids Mfp-3f Mfp-3s Mfp-5 Mfp-6
Pro (P) 6.0 8.0 3.6 4.9
Gly (G) 25.0 29.0 19.6 13.7
Ala (A) 2.0 1.0 2.7 2.9
Cys (C) 0 0 0 2.9
Asp/Asn (D/N) 10.0 18.0 3.6 13.4
Glu (E) 1.0 1.0 0.6 2.3
Ser (S) 1.0 2.0 1.2 4.3
pSer
a
0 0 4.8 2.8
Dopa 19.0 8.0 30.4 3.2
Tyr (Y) 1.0 19.0 0.2 19.2
Trp (W) 6.0 – 0 0
His (H) 1.0 3.0 4.8 0
Lys (K) 15.0 4.0 19.8 9.8
hArg
b
1.0 0 0 0
Arg (R) 9.0 2.0 3.1 6.5
Total 100 99 100 100
Reference 19 19 14 14
a
Phosphoserine.
b
Hydroxyarginine.
Acidic, basic and aromatic residues are shaded blue, red and purple,
respectively
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