Molecular Simulation Study On Two Kinds Of Protein-natural Velvet Antler Polypeptide (nVAP) And Barrier-to-autointegration Factor (BAF)

In this thesis, we applied molecular dynamics (MD) simulation to examine thestructure and functions of natural velvet antler polypeptide (nVAP) andBarrier-to-autointegration factor (BAF). The content mainly includes the followingthree parts:1. Natural velvet antler polypeptide conformation prediction and molecular dockingstudy with TGF-β1complexBased on the chain A structures of hemoglobin (PDB code:1HDS,1IBE,1FAW,3AT5), the three dimensional (3D) structure of natural velvet antler polypeptide(nVAP) was constructed by homology modeling and molecular dynamics (MD)method. The structural rationality was further checked by Profile-3D and Procheck,both of which confirmed that the3D structure of nVAP was reasonable. The modeledstructure indicates that the stable conformation of nVAP is composed of two α-helixes.The extracellular domains of transforming growth factor-β1receptor I (TβRI-ED) andII (TβRII-ED) were docked with nVAP, respectively. The results show that both ofTβR-EDs have high affinity with nVAP which locates near the active center ofTβRII-ED integrating with transforming growth factor-β1(TGF-β1). Otherwise,nVAP can also insert near the “pre-helix extension” of TβRI-ED, which is the keydomain to interact on TGF-β1and TβRII-ED. With the perturbation of nVAP,TβRI-ED can not be recruited by TGF-β1:TβRII-ED complex rigorously. Theintracellular domain of TβRI (TβRI-ID) is not phosphorylated and activated by TβRII.This study shows that nVAP prefers tethering TβRI-ED which is more crucial inTGF-β1:TβRII-ED:TβRI-ED complex. Thus nVAP can disturb the TGF-β1bindingpattern by interacting on TβRs (TβRI and TβRII), further intercepting TGF-β1pathway downstream.2. Molecular simulation investigation on the interaction between barrier-to- autointegration factor or its Gly25Glu mutant and DNAIn order to understand the binding mechanism between Barrier-to-autointegrationfactor (BAF) and DNA, two DNA:BAF complexes with wild type (WT) BAF and itsGly25Glu point mutate type (MT) were generated by molecular docking on the basisof the crystal structures of BAF(PDB code:2ODG, chain A)and DNA (PDB code:2BZF, chain B and C). Then, molecular dynamics (MD) simulations were performedon the two docked structures, as well as BAF(WT) and BAF(MT). The results showthat monomer BAF is more flexible than BAF in DNA:BAF complex, suggesting thatDNA is effective to stabilize conformation of BAF, which is in good agreement withthe experimental results. Besides, the mutated Glu25in DNA:BAF(MT) can changethe BAF conformation to some extent. With deeper investigation on the DNA:BAFstructures, the hydrogen bonds are found to make great contribution to the interactionbetween DNA and BAF. The hydrogen bonds in DNA:BAF(MT)are fewer than thosein DNA:BAF(WT), indicating that the Gly25Glu mutation in BAF has an importanteffect on the hydrogen bonds in the DNA:BAF complex. Besides, the binding freeenergy in DNA:BAF(MT)is also higher than that in DNA:BAF(WT). It results fromthe influence of Glu25side chain on the orientation of Lys6and Lys33in the interfacebetween DNA and BAF. The binding free energy of Lys72, another key residue,decreases a lot in DNA:BAF(MT) anomalously. The decreasing energy causes thedestruction of hydrophobic pocket in the binding site between DNA and BAF(MT).Our results are helpful for further experimental investigations.3. Molecular simulation investigation on the interaction between barrier-to-autointegration factor dimer or its Gly25Glu mutant and LEM domain of emerinWe study the interaction between barrier-to-autointegration factor dimer (BAF2)wild type (WT) or its Gly25Glu mutant type (MT) and LEM domain of emerin(EmLEM). BAF2:EmLEMcomplex was from PDB code:2ODG structure. Nonspecificfragment of double-strand DNA were docked with each chain of BAF2by ZDOCKprogram. The model of DNA2:BAF2:EmLEMwas thus constructed. We mutated eachGly25of BAF2to Glu25using ‘point mutation’ module of Discovery Studio2.5.Then, molecular dynamics (MD) simulations were performed on DNA2:BAF2(WT):EmLEMand DNA2:BAF2(MT):EmLEMcomplexes. The30nsRMSD plots revealed that mutant Glu25could change the whole conformation of MTcomplex. The binding free energy analysis showed that the electronegative residuesAsp57, Glu61and Asp65from chain A, glu36from chain B of BAF2mainlycontribute to interact EmLEM. However, mutated Glu25attenuate the affinity of theamino acid above to EmLEM. Besides, a stable π-π stack between trp62and phe39from chain B in BAF2(WT) is destroyed by Glu25in BAF2(MT). As a result, trp62forms an interaction with glu25, and phe39converts to strengthen the binding affinitywith EmLEM. Besides, Trp62from chain A also forms a stronger interaction with MTGlu25than that of trp62from chain B. Thus, BAF2(MT) has higher affinity withEmLEMthan BAF2(WT). Our results are coordinate with related experiments.