Exposure of protein modified surfaces to air may be necessary in several applications. For example, air contact may be inevitable during the implantation of biomedical devices, for analysis of protein modified surfaces, or for sensor applications. Protein coatings are very sensitive to dehydration and can undergo significant and irreversible alterations of their conformations upon exposure to air. With the use of two compatible solutes from extremophilic bacteria, ectoine and hydroxyectoine, the authors were able to preserve the activity of dried protein monolayers for up to >24 h. The protective effect can be explained by the preferred exclusion model; i.e., the solutes trap a thin water layer around the protein, retaining an aqueous environment and preventing unfolding of the protein. Horseradish peroxidase (HRP) immobilized on compact TiO2 was used as a model system. Structural differences between the compatible solute stabilized and unstabilized protein films, and between different solutes, were analyzed by static time-of-flight secondary ion mass spectrometry (ToF-SIMS). The biological activity difference observed in a colorimetric activity assay was correlated to changes in protein conformation by application of principal component analysis to the static ToF-SIMS data. Additionally, rehydration of the denatured HRP was observed in ToF-SIMS with an exposure of denatured protein coatings to ectoine and hydroxyectoine solutions.

1.
L.
Berrade
,
A. E.
Garcia
, and
J. A.
Camarero
,
Pharm. Res.
28
,
1480
(
2011
).
2.
S.
Hu
,
Z.
Xie
,
J.
Qian
,
S.
Blackshaw
, and
H.
Zhu
,
WIREs Syst. Biol. Med.
3
,
255
(
2011
).
3.
R. A.
Sheldon
and
S.
Pelt
,
Chem. Soc. Rev.
42
,
6223
(
2013
).
4.
A.
Sassolas
,
L. J.
Blum
, and
B. D.
Leca-Bouvier
,
Biotechnol. Adv.
30
,
489
(
2012
).
5.
P. J.
Wang
,
H.
Thissen
, and
P.
Kingshott
,
Acta Biomater.
45
,
31
(
2016
).
6.
D.
Zhou
and
Y.
Ito
,
RSC Adv.
3
,
11095
(
2013
).
7.
S. J.
Stachelek
 et al,
Biomaterials
32
,
4317
(
2011
).
8.
X.
Hu
,
K. G.
Neoh
,
Z.
Shi
,
E.-T.
Kang
,
C.
Poh
, and
W.
Wang
,
Biomaterials
31
,
8854
(
2010
).
9.
J.
Park
,
S.
Bauer
,
A.
Pittrof
,
M. S.
Killian
,
P.
Schmuki
, and
K.
von der Mark
,
Small
8
,
98
(
2012
).
10.
S.
Prestrelski
,
N.
Tedeschi
,
T.
Arakawa
, and
J. F.
Carpenter
,
Biophys. J.
65
,
661
(
1993
).
11.
N.
Xia
,
C. J.
May
,
S. L.
McArthur
, and
D. G.
Castner
,
Langmuir
18
,
4090
(
2002
).
12.
N.
Xia
and
D. G.
Castner
,
J. Biomed. Mat. Res.
67A
,
179
(
2003
).
13.
S. D.
Allison
,
B.
Chang
,
T. W.
Randolph
, and
J. F.
Carpenter
,
Arch. Biochem. Biophys.
365
,
289
(
1999
).
14.
H.
Shi
,
W.-B.
Tsai
,
M. D.
Garrison
,
S.
Ferrari
, and
B. D.
Ratner
,
Nature
398
,
593
(
1999
).
15.
C.
Ha
and
C. B.
Park
,
Biotechnol. Bioeng.
90
,
848
(
2005
).
16.
M.
Tanaka
,
Y.
Machida
,
S.
Niu
,
T.
Ikeda
,
N. R.
Jana
,
H.
Doi
,
M.
Kurosawa
,
M.
Nekooki
, and
N.
Nukina
,
Nat. Med.
10
,
148
(
2004
).
17.
K.
Lippert
and
E. A.
Galinski
,
Appl. Microbiol. Biotechnol.
37
,
61
(
1992
).
18.
S.
Barth
,
M.
Huhn
,
B.
Mattehey
,
A.
Klimka
,
E. A.
Galinski
, and
A.
Engert
,
Appl. Environ. Microbiol.
66
,
1572
(
2000
).
19.
K.
Göller
and
E. A.
Galinski
,
J. Mol. Catal. B Enzym.
7
,
37
(
1999
).
20.
A. D.
Brown
,
Adv. Microb. Physiol.
17
,
181
(
1978
).
21.
S.
Ohtake
,
Y.
Kita
, and
T.
Arakawa
,
Adv. Drug Del. Rev.
63
,
1053
(
2011
).
22.
T.
Arakawa
and
S. N.
Timasheff
,
Biophys. J.
47
,
411
(
1985
).
23.
I.
Yu
,
Y.
Jindo
, and
M.
Nagaoka
,
J. Phys. Chem. B
111
,
10231
(
2007
).
24.
W. F.
Wolkers
,
H.
Oldenhof
, and
B.
Glasmacher
,
Cryobiology
61
,
108
(
2010
).
25.
G.
Lentzen
and
T.
Schwarz
,
Appl. Microbiol. Biotechnol.
72
,
623
(
2006
).
26.
M. S.
Killian
and
P.
Schmuki
,
Surf. Interface Anal.
46
,
193
(
2014
).
27.
J.
Zhang
,
J.
Li
,
T. A.
Craig
,
R.
Kumar
, and
M. L.
Gross
,
Biochemistry
56
,
3523
(
2017
).
28.
J. B.
Lhoest
,
E.
Detrait
,
P. B
Aguilar
, and
P.
Bertrand
,
J. Biomed. Mater. Res.
41
,
95
(
1998
).
29.
C. D.
Tidwell
,
D. G.
Castner
,
S. L.
Golledge
,
B. D.
Ratner
,
K.
Meyer
,
B.
Hagenhoff
, and
A.
Benninghoven
,
Surf. Interface Anal.
31
,
724
(
2001
).
30.
M. S.
Wagner
and
D. G.
Castner
,
Langmuir
17
,
4649
(
2001
).
31.
D. J.
Graham
and
D. G.
Castner
,
Biointerphases
7
,
49
(
2012
).
32.
M.
Cecilio de Oliveira Monteiro
,
P.
Schmuki
, and
M. S.
Killian
,
Langmuir
33
,
13913
(
2017
).
33.
R. N.
Foster
,
E. T.
Harrison
, and
D. G.
Castner
,
Langmuir
32
,
3207
(
2016
).
34.
M. S.
Killian
,
J.-F.
Gnichwitz
,
A.
Hirsch
,
P.
Schmuki
, and
J.
Kunze
,
Langmuir
26
,
3531
(
2010
).
35.
J. C.
Thomas
,
C.
Pacholski
, and
M. J.
Sailor
,
Lab Chip
6
,
782
(
2006
).
37.
J.-B.
Lhoest
,
M. S.
Wagner
,
C. D.
Tidwell
, and
D. G.
Castner
,
J. Biomed. Mater. Res.
57
,
432
(
2001
).
38.
M.
Henry
and
P.
Bertrand
,
Surf. Interface Anal.
41
,
105
(
2009
).
39.
H. E.
Canavan
,
D. J.
Graham
,
X.
Cheng
,
B. D.
Ratner
, and
D. G.
Castner
,
Langmuir
23
,
50
(
2007
).
40.
B. G. M.
Vandeginste
,
D. L.
Massart
,
L. M. C.
Buydens
,
S.
deJong
,
P. J.
Lewi
, and
J.
Smeyers-Verbeke
,
Handbook of Chemometrics and Qualimetrics: Part B
(
Elsevier
,
Amsterdam
,
1998
).
41.
G. I.
Berglund
,
G. H.
Carlsson
,
A. T.
Smith
,
H.
Szöke
,
A.
Henriksen
, and
J.
Hajdu
,
Nature
417
,
463
(
2002
).
42.
A. S.
Rose
and
P. W.
Hildebrand
,
Nucleic Acids Res.
43
,
W576
(
2015
).
43.
M. S.
Killian
,
H. M.
Krebs
, and
P.
Schmuki
,
Langmuir
27
,
7510
(
2011
).
44.
See supplementary material at https://doi.org/10.1116/1.5031189 for S1: Peak lists for positive polarity spectra; S2: solute and protein fragments visibility after coating and assessment of potential obstruction of bioactivity by solute coatings; S3: PCA comparing (hydroxy-)ectoine protected and unprotected HRP (soaking method only); S4: optical images of protected and unprotected samples after solute coating; S5: contact angle measurements of unprotected HRP; and S6: colorimetric assay of enzymatic activity of unprotected samples with solute addition.

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