Colorimetric and electrochemical detection of ligase through ligation reaction-induced streptavidin assembly  

作　　者：Yaliang Huang Ting Sun Wendi Li Lin Liu Gang Liu Xinyao Yi Jianxiu Wang 

机构地区：[1]Hunan Provincial Key Laboratory of Micro&Nano Materials Interface Science,College of Chemistry and Chemical Engineering,Central South University,Changsha 410083,China [2]Henan Province of Key Laboratory of New Optoelectronic Functional Materials,Anyang Normal University,Anyang 455000,China [3]Sanquan College of Xinxiang Medical University,Xinxiang 453003,China

出　　处：《Chinese Chemical Letters》2022年第6期3151-3155,共5页中国化学快报（英文版）

基　　金：the National Natural Science Foundation of China(Nos.22076221,21876208);the Program for Innovative Research Team of Science and Technology in the University of Henan Province(No.21IRTSTHN005);the Hunan Provincial Science and Technology Plan Project,China(No.2019TP1001)。

摘　　要：We propose a concept for ligase detection by conversion of aggregation-based homogeneous analysis into surface-tethered electrochemical assay through streptavidin(SA)-biotin interaction.Sortase A(Srt A)served as the model analyte and two biotinylated peptides(bio-LPETGG and GGGK-bio)were used as the substrates.Srt A-catalyzed ligation of the peptide substrates led to the generation of bio-LPETGGGKbio.The ligation product(bio-LPETGGGK-bio)induced the aggregation and color change of SA-modified gold nanoparticles(Au NPs)through the SA-biotin interactions,which could be assayed by the colorimetric method.Furthermore,we found that the bio-LPETGGGK-bio could trigger the assembly of tetrameric SA proteins with the formation of the(SA-bio-LPETGGGK-bio)nassemblies through the same interactions.The above results were further confirmed by atomic force microscopy and fluorescent imaging.The insulated assemblies were in-situ fabricated at the SA-modified gold electrode,thus hindering the electron transfer of[Fe(CN)_(6)]^(3-/4-) and leading to an increase in the electron-transfer resistance.The capability of the method for the detection of Srt A both in vitro and Staphylococcus aureus(S.aureus)has been demonstrated.Srt A with a concentration down to 1 pmol/L has been determined by the electrochemical analysis,which is lower than that achieved by the colorimetric assay(50 pmol/L).By integrating the advantages of homogeneous reaction and heterogeneous detection,the strategy serves as an ideal means for the fabrication of various sensing platforms by adopting biotin-labeled and sequence-specific peptide or nucleic acid substrates.

关 键 词：LIGASE Electrochemical analysis STREPTAVIDIN Gold nanoparticles Colorimetric assay Electrochemical impedance spectroscopy 

分 类 号：O657[理学—分析化学]