QM/MM Study Of Tungsten-dependent Benzoyl-Coenzyme A Reductase

Enzymes are closely related to the environment,and can be used to degrade pollutants effectively in the environment.Understanding the reaction mechanisms of enzymes plays an important role in the improvement and design of new enzymes for the degradation of pollutants,like aromatic compounds.In recent years,with the advancement of computer science and technology and the development of quantum chemistry,quantum chemical calculations can be used to elucidate the reaction mechanisms of metalloenzymes and to explain the various selectivities.In this thesis,on the basis of the X-ray crystal structure obtained from experiment,molecular dynamics（MD）simulations and quantum mechanics and molecular mechanics（QM/MM）calculations have been used to investigate the reaction mechanism and regioselectivity of the tungsten-dependent benzoyl CoA reductase（BCRs）.The main research contents are as follows:（1）Reaction mechanism of benzoyl-CoA reductaseTwo different models with different QM regions were constructed,potential energy profiles for the reactions in the singlet state and triplet states were calculated,and it was demonstrated that the catalytic reduction of benzoyl-CoA proceeds efficiently only with a protonated His260 residue.The whole reaction includes two consecutive steps.In the first step,the W⁴⁺-bound water molecule delivers a proton to the para carbon C4 of the benzoyl-CoA substrate,coupled with an electron transfer from the W⁴⁺center to the substrate,leading to the formation of a W⁵⁺-radical intermediate.This step was calculated to be rate-limiting,with a barrier of+23.2kcal/mol in the broken-symmetry singlet state at the B3LYP-D3/def2-TZVPP:Charmm level.Subsequently,the protonated His260 residue delivers a second proton to the meta carbon C3（or C5）of the benzoyl-CoA radical,concomitantly with the shift of the second electron from the pyranopterin cofactor rather than the W⁵⁺ion to the substrate,which has a barrier of+19.1 kcal/mol and harvests the cyclohexa-1,5-diene-1-carboxyl-CoA product.The results will help us to understand the dearomatic degradation of other aromatic compounds.（2）Regioselectivity of benzoyl coenzyme A reductaseFor the reduction of benzoyl coenzyme A,not only 3,4-position reduction can take place,but also at the 1,2-position and 2,3-position.By calculating all possible pathways,the 3,4-position reduction was found to have the lowest barrier,mainly due to the binding model of the substrate and also the stability of the radical intermediate.（3）W vs Mo selectivity in benzoyl coenzyme A reductaseConsidering that W and Mo have similar chemical properties,the calculations have been performed to predict the W vs Mo selectivity in Benzoyl Coenzyme A reductase.It was shown that the W is better than Mo in the reduction of the substrate,while Mo is better than W in the oxidation of the substrate.