Analytical Chemistry Chemical Cytometry Quantitates Superoxide
Analytical Chemistry Chemical Cytometry Quantitates Superoxide
Analytical Chemistry Chemical Cytometry Quantitates Superoxide
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Anal. Chem. 2010, 82, 7035–7043<br />
Quantitation, Visualization, and Monitoring of<br />
Conformational Transitions of Human Serum<br />
Albumin by a Tetraphenylethene Derivative with<br />
Aggregation-Induced Emission Characteristics<br />
Yuning Hong, † Chao Feng, ‡ Yong Yu, †,‡ Jianzhao Liu, † Jacky Wing Yip Lam, † Kathy Qian Luo, ‡,§<br />
and Ben Zhong Tang* ,†,#<br />
Nano Science and Technology Program, Department of <strong>Chemistry</strong>, Institute of Molecular Functional Materials,<br />
Bioengineering Program, and Department of <strong>Chemical</strong> and Biomolecular Engineering, The Hong Kong University of<br />
Science and Technology (HKUST), Clear Water Bay, Kowloon, Hong Kong, China, School of <strong>Chemical</strong> and<br />
Biomedical Engineering, Nanyang Technological University, Singapore 637457, and Department of Polymer Science<br />
and Engineering, Institute of Biomedical Macromolecules, Key Laboratory of Macromolecular Synthesis and<br />
Functionalization of the Ministry of Education, Zhejiang University, Hangzhou 310027, China<br />
Human serum albumin (HSA) is a major protein component<br />
of blood plasma, and its assay is of obvious value to<br />
biological research. We, herein, present a readily accessible<br />
fluorescent bioprobe for HSA detection and quantitation.<br />
A nonemissive tetraphenylethene derivative named<br />
sodium 1,2-bis[4-(3-sulfonatopropoxyl)phenyl]-1,2-diphenylethene<br />
(BSPOTPE) is induced to emit by HSA, showing<br />
a novel phenomenon of aggregation-induced emission<br />
(AIE). The AIE bioprobe enjoys a broad working range<br />
(0-100 nM), a low detection limit (down to 1 nM), and a<br />
superior selectivity to albumins. The fluorescent bioassay<br />
is unperturbed by the miscellaneous bioelectrolytes in the<br />
artificial urine. The AIE luminogen can also be used as a<br />
rapid and sensitive protein stain in gel electrophoresis for<br />
HSA visualization. Utilizing the AIE feature of BSPOTPE<br />
and the Förster resonance energy transfer from HSA to<br />
BSPOTPE, the unfolding process of HSA induced by<br />
guanidine hydrochloride is monitored, which reveals a<br />
multistep transition with the involvement of molten globule<br />
intermediates. Computational modeling suggests that<br />
the AIE luminogens dock in the hydrophobic cleft between<br />
subdomains IIA and IIIA of HSA with the aid of hydrophobic<br />
effect, charge neutralization, and hydrogen bonding<br />
interactions, offering mechanistic insight into the<br />
microenvironment inside the hydrophobic cavity.<br />
Human serum albumin (HSA) is the most abundant protein<br />
in the circulatory system and plays multiple biological functions<br />
in the human body. 1 For example, it regulates water balance<br />
between blood and tissues and serves as a physiological carrier<br />
for various endogenous and exogenous substances that are<br />
* To whom correspondence should be addressed. E-mail: tangbenz@ust.hk.<br />
† Nano Science and Technology Program, Department of <strong>Chemistry</strong>, Institute<br />
of Molecular Functional Materials, and Bioengineering Program, HKUST.<br />
‡ Department of <strong>Chemical</strong> and Biomolecular Engineering, HKUST.<br />
§ Nanyang Technological University.<br />
# Zhejiang University.<br />
(1) Peters, T., Jr. Adv. Protein Chem. 1985, 37, 161.<br />
partially soluble in the bloodstream, such as bilirubin, fatty acids,<br />
and drugs. 2-5 The albumin is synthesized in the liver. A low level<br />
of albumin in the blood serum, known as hypoproteinemia, may<br />
indicate liver failure, cirrhosis, and chronic hepatitis. 6 When blood<br />
passes through healthy kidneys, the kidneys filter out waste<br />
products but keep the substances the body needs, such as albumin<br />
and other proteins. Appearance of an excess amount of proteins<br />
in urine is a sign of chronic kidney disease, which may result in<br />
diabetes, high blood pressure, and problems associated with<br />
inflammation in the kidneys. 7 Many studies have been done on<br />
diabetic nephrosis, with microalbuminuria being identified as an<br />
early sign. 8 Microalbuminuria is commonly diagnosed by elevated<br />
protein concentration (30-300 mg/L) in the urine on at least two<br />
occasions. A higher albumin value is regarded as albuminuria.<br />
The urine-testing dipsticks impregnated with pH sensitive dyes<br />
are normally used for daily health monitoring. 9 The level of<br />
albumin caused by microalbuminuria, however, cannot be accurately<br />
assayed by the dipsticks due to their low sensitivity. 10 It<br />
is, thus, of clinical value to develop effective methods for urinary<br />
protein detection and quantitation.<br />
Colorimetric methods, such as Brandford and Lowry assays,<br />
have traditionally been used for protein quantitation in solutions. 11,12<br />
These methods, however, generally lack sensitivity and accuracy,<br />
(2) Carter, D. C.; Ho, J. X. Adv. Protein Chem. 1994, 45, 153.<br />
(3) (a) Neuzil, J.; Stocker, R. J. Biol. Chem. 1994, 269, 16712. (b) Hu, Y. J.;<br />
Liu, Y.; Xiao, X. H. Biomacromolecules 2009, 10, 517.<br />
(4) Berde, C. B.; Hudson, B. S.; Simoni, R. D.; Sklar, L. A. J. Biol. Chem. 1979,<br />
254, 391.<br />
(5) Sjoholm, I.; Ekman, B.; Kober, A. Mol. Pharmacol. 1979, 16, 767.<br />
(6) Murch, S. H.; Winyard, P. J. D.; Koletzko, S.; Wehner, B.; Cheema, H. A.;<br />
Risdon, R. A.; Phillips, A. D.; Meadows, N.; Klein, N. J.; Walker-Smith, J. A.<br />
Lancet 1996, 347, 1299.<br />
(7) Hoogenberg, K.; Sluiter, W. J.; Dullaart, R. P. F. Acta Endrocrinol. 1993,<br />
129, 151.<br />
(8) (a) Mogensen, C. E.; Keane, W. F.; Bennett, P. H.; Jerums, G.; Parving,<br />
H. H.; Passa, P.; Steffes, M. W.; Striker, G. E.; Viberti, G. C. Lancet 1995,<br />
346, 1080. (b) Nomata, S.; Haneda, K.; Moriya, T.; Katayama, S.; Iwamoto,<br />
Y.; Sakai, H.; Tomino, Y.; Matsuo, S.; Asano, Y.; Makino, H. Jpn. J. Nephrol.<br />
2005, 47, 768.<br />
(9) Rose, B. D. Pathophysiology of Renal Disease; McGraw Hill: New York, 1987.<br />
(10) Martinez, A. W.; Phillips, S. T.; Butte, M. J.; Whitesides, G. M. Angew. Chem.,<br />
Int. Ed. 2007, 46, 1318.<br />
10.1021/ac1018028 © 2010 American <strong>Chemical</strong> Society 7035<br />
<strong>Analytical</strong> <strong>Chemistry</strong>, Vol. 82, No. 16, August 15, 2010<br />
Published on Web 07/20/2010