Rational Design And Bioactivity Analysis Of Novel Natiruretic Peptides

Natriuretic peptides are a group of hormones secreted by cardiomyocytes, including ANP,BNP, CNP and other peptides. They have effects on adjacent myocardial cells, vascularsmooth muscle cells through autocrine or paracrine. The binding of NPs with their specificnatriuretic peptide receptor NPRA, NPRB and NPRC, initiate a series of cell signallingprocess,cause natriuresis, diastolic blood vessels, lowering blood pressure, regulate thebalance of electrolyte and other important physiological functions. But the majority ofnatriuretic peptide drugs have poor stability, short half-life, low solubility shortcomings.Therefore, it is necessary to design and develop new type natriuretic peptide drugs with higheractivity and more stability. This paper based on the analysis of NP molecular modelling andbinding mechanism to NP and NPR. Virtual mutation and amino acid by molecular dynamicssimulation were used to design novel drugs with the BNP as template. The biologicalactivities of newly designed NPs were examined by using NPR-NP reporter system. Resultsare shown as follows:Spatial structures of ANP, BNP and CNP were adopted homology modeling techniques,then considered the URO and VNP based on these structures as templates. We found that theN-and C-terminal of ANP, BNP, URO and VNP showed a three-dimensional cross pattern. Itis assumed that this conformation is more convenient for NP binding to the hydrophilic pokectof the receptor.We carried100ns molecular dynamics assays of these5NPs molecules. At unboundstate ANP, CNP and VNP showed more stable and compact structure. The formation ofintramolecular hydrogen bond number is relatively less. We speculate that these may be oneof the reasons for their higher biological activity. These results provide the feasible way forthe design of new NP molecules.Using homology modeling along with the molecular dynamics simulation technique, weestablished the docking model of protein and its ligand was studied. We had successfulconstructed of complex models of NPRA and ANP, BNP, URO, VNP, and homologymodeling of NPRB, built the complex model of NPRB and CNP, VNP. Molecular dynamicssimulations were performed on the100ns system, which analysed the interaction between NP/NPR In NP/NPRA complex, the interaction between BNP and NPRA was stronger, mainlyreflected in the formation of intermolecular hydrogen bond numbers, and the interactionenergy (Interaction energy) is lower. In NP/NPRB composites, the interaction betweenVNP-NPRB is stronger than CNP-NPRB. The stability of the complexes was higher, theinteraction energy is low, the intermolecular hydrogen bond number. This result lays thetheoretic foundation for the new NP molecular design.The virtual mutations of amino acid were carried out based on BNP, and received440single point mutations,1082double point mutations and626three point mutations. Top10mutants with strong stability and lower energy were selected for next procced.After molecular dynamics simulation and analysis of the MD trajectory of the10mutants, two candidate mutant molecule BNPmt1(S20R/S8W/S21R) and BNPmt2 (S19R/G7R/S21R) were identified. Hydrogen bonds formed between NPRA-BNPmt1andNPRA-BNPmt2are53and45pairs, respectively. Interaction energy data showed that theaverage value of BNPmt1is-689.789kcal/mol, BNPmt2-619.348kcal/mol, lower by22.78%and8.82%comparing to the wild type. Simulation results show that, compared withthe wild type BNP, BNPmt2and BNPmt2have higher physiological activity,Suggesting theycould be new drug candidate for the treatment of ADHF.NP activity repoter genes were designed and constructed. The stable cellines whichcarrying NPRA and NPRB were established for high througout screen of NP activities.Biological activity of the design of the BNPmt1and BNPmt2were measured use theconsturcted NRRA repoter platform. BNPmt1and BNPmt2showed higher biological activitythan the wild type BNPwt, which indicated that mutants with high biological activity weresuccess designed.