| 1. |
- Delcey, Mickaël G., 1988-, et al.
(författare)
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Accurate calculations of geometries and singlet-triplet energy differences for active-site models of [NiFe] hydrogenase
- 2014
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Ingår i: Physical Chemistry, Chemical Physics - PCCP. - : Royal Society of Chemistry (RSC). - 1463-9076 .- 1463-9084. ; 16:17, s. 7927-7938
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Tidskriftsartikel (refereegranskat)abstract
- We have studied the geometry and singlet-triplet energy difference of two mono-nuclear Ni2+ models related to the active site in [NiFe] hydrogenase. Multiconfigurational second-order perturbation theory based on a complete active-space wavefunction with an active space of 12 electrons in 12 orbitals, CASPT2(12,12), reproduces experimental bond lengths to within 1 pm. Calculated singlet-triplet energy differences agree with those obtained from coupled-cluster calculations with single, double and (perturbatively treated) triple excitations (CCSD(T)) to within 12 kJ mol(-1). For a bimetallic model of the active site of [NiFe] hydrogenase, the CASPT2(12,12) results were compared with the results obtained with an extended active space of 22 electrons in 22 orbitals. This is so large that we need to use restricted active-space theory (RASPT2). The calculations predict that the singlet state is 48-57 kJ mol(-1) more stable than the triplet state for this model of the Ni-Sl(a) state. However, in the [NiFe] hydrogenase protein, the structure around the Ni ion is far from the square-planar structure preferred by the singlet state. This destabilises the singlet state so that it is only similar to 24 kJ mol(-1) more stable than the triplet state. Finally, we have studied how various density functional theory methods compare to the experimental, CCSD(T), CASPT2, and RASPT2 results. Semi-local functionals predict the best singlet-triplet energy differences, with BP86, TPSS, and PBE giving mean unsigned errors of 12-13 kJ mol(-1) (maximum errors of 25-31 kJ mol(-1)) compared to CCSD(T). For bond lengths, several methods give good results, e. g. TPSS, BP86, and M06, with mean unsigned errors of 2 pm for the bond lengths if relativistic effects are considered.
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| 2. |
- Irani, Mehdi, et al.
(författare)
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Amino Acid Oxidation of Candida antarctica Lipase B Studied by Molecular Dynamics Simulations and Site-Directed Mutagenesis
- 2013
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Ingår i: Biochemistry. - : American Chemical Society (ACS). - 0006-2960 .- 1520-4995. ; 52:7, s. 1280-1289
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Tidskriftsartikel (refereegranskat)abstract
- Molecular dynamics simulations have been performed on lipase B from Candida antarctica (CalB) in its native form and with one or two oxidized residues, either methionine oxidized to methionine sulfoxide, tryptophan oxidized to 5-hydroxytryptophan, or cystine oxidized to a pair of cysteic acid residues. We have analyzed how these oxidations affect the general structure of the protein as well as the local structure around the oxidized amino acid and the active site. The results indicate that the methionine and tryptophan oxidations led to rather restricted changes in the structure, whereas the oxidation of cystines, which also caused cleavage of the cystine S-S linkage, gave rise to larger changes in the protein structure. Only two oxidized residues caused significant changes in the structure of the active site, viz., those of the Cys-22/64 and Cys-216/258 pairs. Site-directed mutagenesis studies were also performed. Two variants showed a behavior similar to that of native CalB,(M83I and M129L), whereas W155Q and M72S had severely decreased specific activity. M83I had a slightly higher thermostability than native CalB. No significant increase in stability toward hydrogen peroxide was observed. The same mutants were also studied by molecular dynamics. Even though no significant increase in stability toward hydrogen peroxide was observed, the results from simulations and site-directed mutagenesis give some clues about the direction of further work on stabilization.
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| 3. |
- Li, Junhao, 1989-
(författare)
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Theoretical Studies of Drug-Metabolizing Cytochrome P450 Enzymes
- 2020
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The family of cytochrome P450 enzymes (P450s) belongs to one of the most important enzyme families in the human body. P450s are involved in the synthesis of endogenous compounds and metabolism of exogenous substances. In mammalian species, drug metabolizing P450s are anchored in the bilayer lipid membrane, which allows the enzymes to interact with other proteins and ligand molecules. A wealth of knowledge about the structures, functions, and mechanisms of P450s have been obtained from both experimental and theoretical studies. However, the mechanisms behind some experimental results, such as the regio- and stereoselectivity and structural flexibility are still elusive.In this thesis, I present the work done in my doctoral studies, which was focused on the catalytic selectivity and structural flexibility of P450s. Multiple theoretical modeling approaches, such as homology modeling, molecular docking, molecular dynamics, quantum mechanics, and quantum mechanics/molecular mechanics, were applied in the studies. In papers I and II, the regio- and stereoselectivity of CYP4F2, CYP3A4, and CYP19A1 catalyzed C–H hydroxylation of different substrates were studied. The results indicate that the ligand reactivity and accessibility can be decisive for the regio- and stereoselectivity. However, which of them is more important is system-dependent. The quantum mechanics/molecular mechanics calculation results imply that the distribution of spin natural orbitals could be used for discriminating the roles of the reactivity and accessibility. In papers III and IV, the conformational dynamics of the open and closed structures of CYP2B4 and the ligand cooperativity phenomenon of midazolam metabolized by CYP3A4 were investigated using molecular dynamics simulations. From the simulation results, we identified the key residues for the conformational dynamics for the open-to-intermediate transition and found that the ligand cooperativity is also caused by the large flexibility of P450. The results also indicated that the homotropic cooperativity mainly occurs in the large and flexible productive site, rather than in the remote allosteric site.
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