Screening Of Fungal Immunomodulatory Protein And Its Biological Activity

Fungal immunomodulatory proteins(Fungal immunomodulatory protein,FIP)is a kind of structure and function and plant lectins and immunoglobulin heavy chain variable region,has the agglutination of red blood cells,promote human peripheral blood lymphocyte proliferation and stimulate the immune cells to produce a variety of cytokines immunoregulatory activity of small molecules such as proteins.Due to its high immune activity,low effective concentration,simple composition and easy purification,it has good clinical application prospect and medicinal health value.Although various fungal immunoregulatory proteins have certain similarities in primary structure,secondary structure and advanced structure,they have similar biological functions.However,different FIPs perform differently in certain biological activities.Therefore,dig new fungal immunomodulatory proteins in the fungus,to explore more biological function or higher immune regulating the activity of medicinal proteins provide rich biological resources,and for its development as a new type of drug or functional foods provide valuable theoretical foundation and application basis.This article with ganoderma lucidum immunomodulatory protein LZ-8 as the template,the straw mushrooms(Volvaria volvacea)genome of BLAST found with LZ similarity in more than 60% of 7-8 kinds of FIP,except the FIP-vvo97 as straw mushroom immune regulatory proteins have been reported,the other 6 kinds of FIP are new type of fungal immunomodulatory proteins,we will be named the FIP-vvo77 respectively,FIP-vvo78,FIP-vvo79,FIP-vvo80,FIP-vvo82,and FIP-vvo98.First of all,the methods of bioinformatics,will these new FIP with known fungal immunomodulatory proteins in physiological and biochemical properties,the subcellular localization,signal peptide,compares and analyses the genetic evolution,etc,to predict the activity of new FIP.Then,building engineering bacteria in vitro,induction of new FIP protein expression and purification,get the target protein,testing new immunoregulatory activity of FIP and promote cell apoptosis activity,based on molecular level and research a new kind of FIP Hela cell apoptosis pathway,A549 and Spca-1,the influence of the screening bioactive better FIP,development and application for subsequent studies provide important theoretical basis.The specific results are as follows:1.7 new fungal immunomodulatory proteins are composed of about 113 amino acid residues,free of cysteine(Cys),rich in valine(Val)and Tyr.The relative molecular weight is 12-13 k D,and the theoretical equivalence point p I is from 6.06 to 8.83.Through comparison of Fip-vvo82 unstable factor(II)the highest(20.77),it Fip-vvo82 compared with other new fungal immunomodulatory proteins,flexibility is good,may have a highly active,and through the molecular simulation method to predict Fip-vvo82 produce highly active molecular mechanism.2.Test the new type of straw mushroom immune regulatory proteins stimulate immune cells secrete interleukin-2,the experimental results are consistent with the raw letter,namely the new fungal immunomodulatory proteins FIP-vvo82 immune activity.3.6 new fungal immunomodulatory proteins(fip-vvo78,79,80,82,98,and fip-vvo)affect the vitality and apoptosis of Hela,A549 and Spca-1 cells.FIP-vvo82 to Hela cells and promote apoptosis of A549 and Spca-1 effect is more obvious,which promote apoptosis of Hela and A549 cells effect is most obvious,apoptosis rate were 63.4% and 54.8% respectively,the FIP-vvo-79,FIP and FIP-vvo vvo-80-97 the Spca-1 cell and promote apoptosis effect is significant,the apoptosis rate is respectively: 73.4%,73% and 71.3%.4.The effects of 6 new fungal immunoregulation proteins(fip-vvo78,79,80,82,98,and fip-vvo)on the apoptotic signaling pathway of Hela,A549 and Spca-1 cells.Seven kinds of new type of fungal immunomodulatory proteins by blocking the generation of mitochondrial membrane potential(MMP),reduce the apoptosis related proteins the Bcl-2 and the Bcl-xl,improve the activity of Caspase 3,improve the cytochrome C(Cyto C)released from mitochondria intensity,thus promotes the apoptosis of tumor cells.