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| Construction and Application of a Functional Peptide-mediated Fluorescence Biosensor for Copper Ions Detection |
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| DOI： |
| KeyWord:peptides copper ions metalloprotein fluorescence detection |
| Author | Institution |
| LI Xin-yi,FENG Fu-yan，ZHOU Xiao-dong,HU Ji-ming |
The Centre of Analysis and Measurement of Wuhan University,Wuhan University,Wuhan,China |
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| Abstract: |
| Problems of heavy-metal pollution have been drawing more and more attraction of all parts of society.To establish a simple,rapid,stable and efficient detection method for heavy metal ions is an urgent need for environmental stability and life-health.The copper ions(Cu(Ⅱ)) may be an important factor leading to several diseases,including Alzheimer's disease,prion disease and amyotrophic lateral sclerosis.Although some significant researches have been reported in development of chemosensors to detect Cu(Ⅱ),the development of peptide-based fluorescent chemosensors capable of detecting multiple ions has received considerably less attention.The metalloprotein database offers a great possibility for the design and screening of sensing elements in metal ions detection.Through the comparative analysis of the interactions between biomolecules(e-g.proteins,peptides and amino acids) and metal ions,the high specificity and stability of the binding of peptide molecules to metal ions have been determined.In combination with such advantages of peptide molecules as stability,flexibility,and easy modification,a series of analytical studies were conducted on their behaviors with metal ions.This paper was devoted to the construction of a fluorescent Cu(Ⅱ) biosensor based on the specific performance between peptides and Cu(Ⅱ) ions.In view of the flexible design and coordination sites of peptides,the interaction between peptides and Cu(Ⅱ) was deeply researched.A peptide(DDAEGHARHCR) specifically coordinated with Cu2+ was engineered by utilizing the known amyloid protein.And their binding ability and specifity were confirmed by the characterization of MALDI-TOF MS.With the formation of the ternary-competition system “peptides-Cu(Ⅱ)-calcein”,a unique fluorescence biosensor was constructed,thereby achieving the highly sensitive detection of Cu(Ⅱ) in water samples.As is known,Cu(Ⅱ) ions could effectively quench the fluorescence signal of calcein.When the functional peptide was introduced into the calcein-Cu(Ⅱ) system,the peptide with higher binding ability could promptly seize the Cu(Ⅱ) ions,greatly recovering the fluorescence signal of this system.Thus,the effective Cu(Ⅱ) sensing detection could be built.After optimizing the reaction temperature,reaction time,pH and calcein concentration,the detection of Cu(Ⅱ) in environmental water samples was also realized.Results demonstrated that the detection limit of this method for Cu(Ⅱ) was as low as 127 nmol/L,with a linear range of 0.12-13 μmol/L.To ensure the reliabity,the Cu(Ⅱ) concentrations was quantified by atomic absorption spectroscopy(AAS).The high consistency between AAS and our results indicated the accuracy and dependability of the work.Compared with other approaches for Cu2+ detection,the established method exhibited superiorities in simplicity,selectivity,efficiency and stability.Furthermore,the protocol involving the peptides design had a great application potential and a broad development prospect in pratical detection,which also offered a novel idea for the constitution of recognition elements. |
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