Study On The Conformational Change Of The Flocculation Protein Flo1 Active Site

The interaction between flocculation protein and the chain of polysaccharide leads to the flocculation.Flo1 p is the most dominant flocculation protein.N-Flo1 p is the N terminal of Flo1 p and existing in the surface of Saccharomyces cerevisiae interacts with the mannosan on the cell wall.Exploring the interaction between flocculation protein-ligand contribute to the research on the flocculation.There are two known conformer of N-Flo1 p.One is the unbound state(PDB ID: 4LHL),another is the bound state with mannose(PDB ID: 4LHN).The binding site of 4LHN is obviously narrower than that in 4LHL,indicating mannose induced the binding site of NFlo1 p shrinking.Understanding the change process of N-Flo1 p binding pocket is vital to build up the flocculation model.However,obtaining the transition path of protein conformation change is challenging,because of the complicated transition mechanism and hige degree of freedom.The research of this article is centerd on protein conformation changing process and the accompanying influences on the protein function are discussed in this article.1.We applied different methods to produce the intermediate states of protein.The results demonstated that Cartesian coordinate interpolation could not be applied to the flexbile sidechain motion(such as aromatic ring overturn);Backbone interpolation-sidechain transposition could not lead to sidechain internal structure change;Though internal coordinate interpolation could lead to the internal structure change,backbone crossover exsists in the intermediate state.2.Presenting a combined interpolation method.After knowing the advantages and disadvantages of different methods,we proposed BISO(Backbone Interpolation Sidechain Orientation)method.Specifically the method assembling the linear interpolated backbone and the internal coordinate interpolation sidechain.Analysising the intermediate state,we found BISO method could not lead to backbone crossover and sidechain transformation.In the following structural optimization,to eliminate the unreasonable atomic interaction of intermediate structure,we designed the minimization schemes which could respectively be suitable for backbone and sidechain.3.The molecular dynamics simulation of the two conformation.We used AMBER14 software to run molecular dynamics of the two complexes and found that mannose could only stay in the binding site 6 ns,after 6 ns mannose run away from the binding site.However,for 4LHN mannose stable stayed in the binding site of 4LHN for 50 ns.The binding free energy calculation results shown that calcium ion played an important role in stabilizing mannose;Asp160,ASP161 interact strongly with ligand when mannose entering the binding pocket;The shrinkage of binding site strengthen the interaction between mannose with LYS194 and GLN94.We wish BISO method could be able to provide the initial path of protein conformation change and offer help in finding conformation transition path of the macromolecular protein system.