Tertiary Structure Modelling And Dimerization Study Of The FNR Protein From Klebsiella Pneumoniae

Klebsiella pneumoniae is a metabolically versatile bacterium that is able togrow in both the presence and absence of oxygen. It can be used for producingchemicals such as1,3-propanediol,2,3-buanediol, etc. under anaerobic and aerobicconditions by its own oxygen regulatory system. Its oxygen regulation mechanismhas attracted increasing interest. FNR (fumarate nitrate reduction regulator) is animportant oxygen regulating protein in K. pneumoniae. Its activity varies accordingto oxygen concentrations. FNR appears to sense oxygen directly through aredox-sensitive iron-sulphur cluster in the protein. It exists in active dimer form onlyduring anaerobic growth. Active FNR protein activates and represses target genes bybinding with the specific DNA in response to anaerobic conditions. Upon exposureto oxygen, the FNR doesn’t bind to the specific DNA and the activity of FNR alsodisappears, additionally, the dimer form converts into monomer form. At present, theoxygen regulation mechanism of FNR is not very chear. The modelling of its tertiarystructure and the analyses of its dimerization is helpful for studying its oxygenregulatory mechanism.Firstly, the tertiary structure of FNR monomer in K. pneumoniae wasconstructed by homology modelling, based on the CRP and endonuclease Ⅲ as thetemplates. In addition, it was further optimized and processed by molecularmechanics and molecular dynamics simulation. The structure rationality evaluatedthrough PROCHECK, ERRAT, Verify-3D and PROSA programs indicated that theFNR protein monomer structure was reasonable.Next, the interface residues of FNR monomer were predicted by PPI-Pred server combined with interface analysis. The FNR dimer was obtained by dockingtwo monomers, and the dimer structure was further optimized to be the bestconformation through optimization and dynamics simulation.At last, the electrostatic potential interaction, the hydrophobic interaction, thehydrogen bond interaction and the interaction energy analyses were conducted. Itwas revealed that hydrophobic interactions and hydrogen bonding were the mainfactors for driving the stability of FNR dimer. The Robetta server was used foranalyzing the alanine mutation effect. It was found that Arg140, Met144, Arg145,Met147, Glu150, Ile151and Ile158were hot residues, which could promote thedimerization of the FNR dimer in anaerobic conditions. However, Gln141, Ile152,Gln155, Met157, Leu159, Leu161and Ser162had small effect on the stability of theFNR dimer. In addition, Ser148and Asp154would inhibit the dimerization of theFNR dimer.