Studies On The Application Of Anti-phosphoarginine Antibody In Proteome And Protective Effect Of Montmorillonite On Peptides Under Prebiotic Conditions

Reversible phosphorylation of proteins plays an important role in vivo.As a kind of protein phosphorylation,N-phosphorylation is often neglected due to some special reasons,such as acid instability,low natural abundance and lack of effective research tools.Based on this,a magnetic nanoparticles with gold nanoparticles modified on the surface(Fe3O4@SiO2@Au)was used as a pArg antibody immobilization tool to study one of the N-phosphorylation-arginine phosphorylation.The purified pArg antibody was connected to Fe3O4@SiO2@Au and we used it as enrichment tools to explore new methods for pArg enrichment.The change of Zeta potential value and SDS-PAGE result indicate that the pArg antibody could successfully attach to Fe3O4@SiO2@Au,and still remains biological activity.Using this enrichment tool,we successfully enriched the McsB protein-a protein that contained the pArg site.In addition,considering the instability of N-phosphorylation,we optimized the mobile phase condition of nano LC-MS and increased the pH value of the mobile phase to 4.McsB was used to verify the detection effect of nano LC-MS.7 pArg sites on 6 peptides of McsB protein were successfully identified.The results show that the optimized nano LC-MS detection condition has a good detection effect for McsB.In the process of chemical evolution,the stability of prebiotic organic compounds is very important,especially in the presence of ultravioiet(UV)radiation,which would cause massive destruction of prebiotic organic compounds.Montmorillonite(MMT),a clay that may have existed in large quantities on primitive earth,has been proved to be a good catalyst and adsorbent,playing an important role in the chemical evolution of biological molecules.Herein,the ability of MMT to absorb peptides at pH 3 and 10 and its ability to resist UV radiation after adsorption were systematically studied.Our research indicates that the adsorption of Phe,Phe2,Tyr2,Val2,Ala2,Leu2,Pro2,FFFFD from aqueous solution to MMT is affected by pH condition.Acidic condition(pH=3)is beneficial to the adsorption of peptides on MMT.Moreover,the adsorption rate of peptides on MMT was greater than 60%.While basic condition(pH=10)is good for the desorption from MMT.In addition,by comparing the adsorption rates of Phe,Phe2 and FFFFD on MMT,it can be seen that the adsorption capacity of MMT is proportional to the length of the peptide chain.More importantly,we first discovered that using the adsorption of MMT at pH 3 can better help the peptides resist the decomposition of UV radiation.Compared with other peptides,MMT has more significant protective effect on Phe2 at pH 3.And combined with previous research results,a new possible protective mechanism of MMT on peptides was proposed.That is,peptides can be adsorbed into the interlayer of MMT through electrostatic adsorption and ion exchange,which reduces the UV radiation intensity of peptides and thus better protects peptides from UV decomposition.