Exploration On Composition And Properties Of The Low Molecular Weight Fluorescent Stripes From Anabaena Sp.PCC7120by Electrophoresis Separation

Fluorescent protein is considered as a revolution in biology research. Thespectrum of fluorescent proteins such as GFP that had been reported almost cover theentire visible region, which are widely used as one of the most significant tools inmodern biological research, being generally applied in gene expression regulation,protein localization and translation, protein folding, signal transduction, enzymaticassay，biomolecular interaction and molecular imaging. Phycobilisomes are part ofrepresentative fluorescent protein from alga. For the advantage of high quantum yieldand stability, it has application fields in biological research such as FACS, FIA andSMD.This study made electrophoresis analysis about fluorescent substance fromAnabaena sp.PCC7120, and found some fluorescent stripes those have differentmolecular weight from known fluorescent substance such as phycobilisome subunits，chlorophyll-protein complexes and free chlorophyll. By the research of compositionand properties of fluorescent stripes, we may provide new material of fluorescent tagand fluorescent probe. Also by the comparison of this fluorescent substance withphycobilisome subunits and chlorophyll-protein complexes，we may discover thesource and possible function about these fluorescent substance.This research is an exploration based on the facts below. These fluorescentstripes have low molecular weight less than10kD， high temperatureresistance, high quantum yield and long wave emission peak, which have possibilityusing as new fluorescent tag or fluorescent probe with highly application future.This research is mainly consist of the following contents. Separated proteinsfrom Anabaena sp.PCC7120by electrophoresis, compared with those knownfluorescent stripes and marked those unknown. Meanwhile, we studied theconnections between different fluorescent stripes by ways such as ultrasonicprocessing, high-temperature processing and than going a secondary electrophoresis.For the unknown fluorescent stripes which are less than10kD, an experiment wasdesigned including2-D gel electrophoresis and MALDI-TOF to analyze the possiblecomposition, than formed a hypothesis and finally confirm this by molecular cloning.Moreover, we analyzed the pigment part of these fluorescent substance by spectralanalysis, ICP-AES and florescence reaction that based on the special combining ability between Zn2+and porphyrin. Next，we analyzed the protein part of thesefluorescent substance by mass spectrometric analysis， BCA and ninhydrin proteinassay. Finally, we made a general judgment on composition and function of thesefluorescent stripes，which would help to show its capability in being used asfluorescent tag.After a series of experiments, we got some preliminary experiment results. Thepigment may has porphyrin ring which is similar with Chlorophyll, phycobilin andheme. As we known, free proline has important effections to the plant physiologicalregulation, especially resistance abilities, while photosynthetic pigments such asChlorophyll and phycobilin are necessary component for photosynthesis. Thereforethe analysis of these fluorescent stripes will have the possibility not only using as newfluorescent tag or fluorescent probe, but also being helpful to the function andregulatory mechanism of proline by combining with this two important components.On account of the preliminary results only at present, we still have a long way to goon our research.