The Design And Synthesis Of Fluorescent Probes Based On Peptide Chemical Reaction And The Biological Applications

In the field of life sciences,in-depth exploration of protein chemistry is one of the important means to understand life processes.In the cell,the main chain and the residues of the protein can participate in different chemical reactions,such as coordination,acetylation,methylation,glycosylation and so on.These chemical reactions play an important role in molecular recognition,nerve signal transduction and metabolism.Further study of the relationship between these chemical reactions and biological signal conduction is a significant basis for revealing the life process.Since solid-phase peptide synthesis won the Nobel Prize in Chemistry,the artificial design and synthesis of peptide molecules has been realized.This contribution laid the foundation for intensive research of the chemical reactions involved in proteins.The peptide contains a large number of precisely located residues and different functional groups on its side chains can participate in chemical reactions of target molecules both in vitro and in vivo.In addition,the affinity and specificity of the target analyte can be improved.The specific peptide sequences linked fluorophores is a common strategy for designing peptide fluorescent probes.In the process of interacting with the analyte,the fluorophore acts as a signal output to obtain relevant chemical reaction information.Compared with fluorescent proteins,peptide fluorescent probes are more stable and can be easily modified at specific sites in biological applications.Therefore,different peptide sequences and response mechanisms can be designed for bioimaging and disease marker detection.According to the various advantages of peptide fluorescent probes,the main studies are as follows:1)We chose fluorescein isothiocyanate as the fluorophore.By connecting the peptide backbone and considering the type and quantity of amino acids contained in the peptide,we successfully designed the peptide fluorescent probe FP.When FP is highly selectively coordinated with Cu²⁺,FP-Cu²⁺can be used to detect glutathione.This first developed peptide fluorescent probe for detecting GSH has the advantage of instant response and has been successfully used for the qualitative and quantitative detection of Cu²⁺and GSH.At the same time,this method can be applied to live cell and zebrafish imaging,which has potential value for biological applications.2)We studied the effects of amino acids and peptides on the coordination ability of metal ions.Through subtle adjustments to the structure of the peptide,a variety of analytes are recognized.The two polypeptide fluorescent probes have the same fluorophore and linker,and each responds to different metal ions due to the difference in coordination groups.By comparing the properties of the two peptide probes,it is proved that KEEP can show the only response to Hg²⁺from the"soft acid"metal ion.This phenomenon is very rare.In addition,the opposite fluorescence response of KEEP to Hg²⁺and CH₃Hg⁺also provides a new idea for the future design of fluorescent probes that can effectively distinguish inorganic mercury from organic mercury.3)Peptide fluorescent probe TP-2 designed based on the aggregation-induced emission effect can coordinate with Hg²⁺with high selectivity.After coordination,the aggregation and intramolecular rotation of TPE derivative fluorescent molecules are restricted,which increases the fluorescence emission signal of TP-2 by about 30 times.High signal-to-noise ratio and high sensitivity are obvious advantages.In cell imaging,TP-2 and cytosolic lysosomal dye co-stain cells to achieve a high degree of co-localization.TP-2 has excellent cell membrane penetration and cell permeability,and has potential value in further bioimaging applications.4)Based on the aggregation-inducing effect andβ-turn structure,the peptide fluorescent probe TP was designed.The coordination ratio between TP and Cd²⁺is 2:1.The highly selective coordination shortens the distance between the fluorophores,leading to the aggregation of TP and strong fluorescence emission.In addition,without changing the structure of the connecting partβ-turn,the specific coordination of Zn²⁺is achieved by optimizing the types of amino acids with coordination ability.We successfully transferred the detection system to the test strip,and realized the qualitative and quantitative detection of Cd²⁺by analyzing the fluorescence enhancement efficiency on the test strip.The test paper strip is convenient to carry,low in cost,and high in stability,and can be applied to the actual detection of biological and environmental samples.At the same time,it provides a potential direction for the design of low-toxic heavy metal ion chelating agents.