Controlling the Substrate Specificity of an Enzyme through Structural Flexibility by Varying the Salt-Bridge Density

Juan Huang, Qin Xu, Zhuo Liu, Nitin Jain, Madhusudan Tyagi, Dong-Qing Wei, Liang Hong
2021 Molecules  
Many enzymes, particularly in one single family, with highly conserved structures and folds exhibit rather distinct substrate specificities. The underlying mechanism remains elusive, the resolution of which is of great importance for biochemistry, biophysics, and bioengineering. Here, we performed a neutron scattering experiment and molecular dynamics (MD) simulations on two structurally similar CYP450 proteins; CYP101 primarily catalyzes one type of ligands, then CYP2C9 can catalyze a large
more » ... ge of substrates. We demonstrated that it is the high density of salt bridges in CYP101 that reduces its structural flexibility, which controls the ligand access channel and the fluctuation of the catalytic pocket, thus restricting its selection on substrates. Moreover, we performed MD simulations on 146 different kinds of CYP450 proteins, spanning distinct biological categories including Fungi, Archaea, Bacteria, Protista, Animalia, and Plantae, and found the above mechanism generally valid. We demonstrated that, by fine changes of chemistry (salt-bridge density), the CYP450 superfamily can vary the structural flexibility of its member proteins among different biological categories, and thus differentiate their substrate specificities to meet the specific biological needs. As this mechanism is well-controllable and easy to be implemented, we expect it to be generally applicable in future enzymatic engineering to develop proteins of desired substrate specificities.
doi:10.3390/molecules26185693 pmid:34577164 pmcid:PMC8470667 fatcat:b5jheekkfjddtefyjiclh7curu