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Siegbahn, P. E. M. (2024). Computational Model Study of the Experimentally Suggested Mechanism for Nitrogenase. Journal of Physical Chemistry B, 128(4), 985-989
Open this publication in new window or tab >>Computational Model Study of the Experimentally Suggested Mechanism for Nitrogenase
2024 (English)In: Journal of Physical Chemistry B, ISSN 1520-6106, E-ISSN 1520-5207, Vol. 128, no 4, p. 985-989Article in journal (Refereed) Published
Abstract [en]

The mechanism for N-2 activation in the E-4 state of nitrogenase was investigated by model calculations. In the experimentally suggested mechanism, the E-4 state is obtained after four reductions to the ground state. In a recent theoretical study, results for a different mechanism have been found in excellent agreement with available Electron Paramagnetic Resonance (EPR) experiments for E-4. The two hydrides in E-4 leave as H-2 concertedly with the binding of N-2. The mechanism suggested differs from the experimentally suggested one by a requirement for four activation steps prior to catalysis. In the present study, the experimentally suggested mechanism is studied using the same methods as those used in the previous study on the theoretical mechanism. The computed results make it very unlikely that a structure obtained after four reductions from the ground state has two hydrides, and the experimentally suggested mechanism does therefore not agree with the EPR experiments for E-4. Another structure with only one hydride is here suggested to be the one that has been observed to bind N-2 after only four reductions of the ground state.

National Category
Organic Chemistry
Identifiers
urn:nbn:se:su:diva-226560 (URN)10.1021/acs.jpcb.3c07675 (DOI)001156037200001 ()38237063 (PubMedID)2-s2.0-85183498894 (Scopus ID)
Available from: 2024-02-14 Created: 2024-02-14 Last updated: 2024-02-14Bibliographically approved
Siegbahn, P. E. M. (2024). Mechanisms for Methane and Ammonia Oxidation by Particulate Methane Monooxygenase. Journal of Physical Chemistry B
Open this publication in new window or tab >>Mechanisms for Methane and Ammonia Oxidation by Particulate Methane Monooxygenase
2024 (English)In: Journal of Physical Chemistry B, ISSN 1520-6106, E-ISSN 1520-5207Article in journal (Refereed) Epub ahead of print
Abstract [en]

Particulate MMO (pMMO) catalyzes the oxidation of methane to methanol and also ammonia to hydroxylamine. Experimental characterization of the active site has been very difficult partly because the enzyme is membrane-bound. However, recently, there has been major progress mainly through the use of cryogenic electron microscopy (cryoEM). Electron paramagnetic resonance (EPR) and X-ray spectroscopy have also been employed. Surprisingly, the active site has only one copper. There are two histidine ligands and one asparagine ligand, and the active site is surrounded by phenyl alanines but no charged amino acids in the close surrounding. The present study is the first quantum chemical study using a model of that active site (Cu-D). Low barrier mechanisms have been found, where an important part is that there are two initial proton-coupled electron transfer steps to a bound O-2 ligand before the substrate enters. Surprisingly, this leads to large radical character for the oxygens even though they are protonated. That result is very important for the ability to accept a proton from the substrates. Methods have been used which have been thoroughly tested for redox enzyme mechanisms.

National Category
Biochemistry and Molecular Biology
Identifiers
urn:nbn:se:su:diva-231265 (URN)10.1021/acs.jpcb.4c01807 (DOI)001242686700001 ()38850249 (PubMedID)2-s2.0-85195558363 (Scopus ID)
Available from: 2024-06-19 Created: 2024-06-19 Last updated: 2024-06-19
Siegbahn, P. E. M. & Wei, W.-J. (2024). The energetics of N2 reduction by vanadium containing nitrogenase. Physical Chemistry, Chemical Physics - PCCP, 26(3), 1684-1695
Open this publication in new window or tab >>The energetics of N2 reduction by vanadium containing nitrogenase
2024 (English)In: Physical Chemistry, Chemical Physics - PCCP, ISSN 1463-9076, E-ISSN 1463-9084, Vol. 26, no 3, p. 1684-1695Article in journal (Refereed) Published
Abstract [en]

The main class of nitrogenases has a molybdenum in its cofactor. A mechanism for Mo-nitrogenase has recently been described. In the present study, another class of nitrogenases has been studied, the one with a vanadium instead of a molybdenum in its cofactor. It is generally believed that these classes use the same general mechanism to activate nitrogen. The same methodology has been used here as the one used for Mo-nitrogenase. N2 activation is known to occur after four reductions in the catalytic cycle, in the E4 state. The main features of the mechanism for Mo-nitrogenase found in the previous study are an activation process in four steps prior to catalysis, the release of a sulfide during the activation steps and the formation of H2 from two hydrides in E4, just before N2 is activated. The same features have been found here for V-nitrogenase. A difference is that five steps are needed in the activation process, which explains why the ground state of V-nitrogenase is a triplet (even number) and the one for Mo-nitrogenase is a quartet (odd number). The reason an additional step is needed for V-nitrogenase is that V3+ can be reduced to V2+, in contrast to the case for Mo3+ in Mo-nitrogenase. The fact that V3+ is Jahn–Teller active has important consequences. N2H2 is formed in E4 with reasonably small barriers.

National Category
Inorganic Chemistry
Identifiers
urn:nbn:se:su:diva-225645 (URN)10.1039/d3cp04698b (DOI)001130246900001 ()38126534 (PubMedID)2-s2.0-85180595702 (Scopus ID)
Available from: 2024-01-31 Created: 2024-01-31 Last updated: 2024-03-11Bibliographically approved
Siegbahn, P. E. M. (2023). Can the E1 state in nitrogenase tell if there is an activation process prior to catalysis?. Physical Chemistry, Chemical Physics - PCCP, 25(5), 3702-3706
Open this publication in new window or tab >>Can the E1 state in nitrogenase tell if there is an activation process prior to catalysis?
2023 (English)In: Physical Chemistry, Chemical Physics - PCCP, ISSN 1463-9076, E-ISSN 1463-9084, Vol. 25, no 5, p. 3702-3706Article in journal (Refereed) Published
Abstract [en]

Model calculations have been performed for the singly reduced ground state of Mo-nitrogenase, usually termed E1. Contradictory conclusions have been reached in two recent experimental studies. In a study based on EPR, it was concluded that there is a bridging hydride in E1, while in an X-ray study it was concluded that there is no hydride in E1. Therefore, the EPR study implies that there is an oxidation of the cofactor going from E0 to E1, the X-ray study implies a reduction. DFT methods have here been used, which have previously been benchmarked on a set of redox enzymes that led to the conclusion that the accuracy is about 3 kcal mol−1 in all cases, even for redox transitions. The methodology should therefore be adequate for resolving the question of the hydride presence in E1. As a comparison, calculations are performed on both Mo- and V-nitrogenase with the same conclusion. The conclusion from the calculations has far reaching consequences for the mechanism of nitrogenase.

National Category
Chemical Sciences
Identifiers
urn:nbn:se:su:diva-214811 (URN)10.1039/d2cp05642a (DOI)000915305400001 ()36655689 (PubMedID)2-s2.0-85146827471 (Scopus ID)
Available from: 2023-02-15 Created: 2023-02-15 Last updated: 2023-02-15Bibliographically approved
Siegbahn, P. E. M. (2023). Computational modeling of redox enzymes. FEBS Letters, 597(1), 38-44
Open this publication in new window or tab >>Computational modeling of redox enzymes
2023 (English)In: FEBS Letters, ISSN 0014-5793, E-ISSN 1873-3468, Vol. 597, no 1, p. 38-44Article, review/survey (Refereed) Published
Abstract [en]

A computational methodology is briefly described, which appears to be able to accurately describe the mechanisms of redox active enzymes. The method is built on hybrid density functional theory where the inclusion of a fraction of exact exchange is critical. Two examples of where the methodology has been applied are described. The first example is the mechanism for water oxidation in photosystem II, and the second one is the mechanism for N2 activation by nitrogenase. The mechanism for PSII has obtained very strong support from subsequent experiments. For nitrogenase, the calculations suggest that there should be an activation process prior to catalysis, which is still strongly debated. 

Keywords
hybrid density functional theory, mechanisms, nitrogenase, photosystem II, redox enzymes
National Category
Chemical Sciences
Identifiers
urn:nbn:se:su:diva-212484 (URN)10.1002/1873-3468.14512 (DOI)000871607800001 ()36254111 (PubMedID)2-s2.0-85140385502 (Scopus ID)
Available from: 2022-12-08 Created: 2022-12-08 Last updated: 2024-03-26Bibliographically approved
Siegbahn, P. E. M. (2023). How Protons Move in Enzymes - The Case of Nitrogenase. Journal of Physical Chemistry B, 127(10), 2156-2159
Open this publication in new window or tab >>How Protons Move in Enzymes - The Case of Nitrogenase
2023 (English)In: Journal of Physical Chemistry B, ISSN 1520-6106, E-ISSN 1520-5207, Vol. 127, no 10, p. 2156-2159Article in journal (Refereed) Published
Abstract [en]

When moving protons in enzymes, water molecules are often used as intermediates. The water molecules used are not necessarily seen in the crystal structures if they move around at high rates. In a different situation, for metal containing cofactors in enzymes, it is sometimes necessary to move protons on the cofactor from the position they enter the cofactor to another position where the energy is lower. That is, for example, the situation in nitrogenase. In recent studies on that enzyme, prohibitively high barriers were sometimes found for transferring protons, and that was used as a strong argument against mechanisms where a sulfide is lost in the mechanism. A high barrier could be due to nonoptimal distances and angles at the transition state. In the present study, possibilities are investigated to use water molecules to reduce these barriers. The study is very general and could have been done for many other enzymes. The effect of water was found to be very large in the case of nitrogenase with a lowering of one barrier from 15.6 kcal/mol down to essentially zero. It is concluded that the effect of water molecules must be taken into account for meaningful results.

Keywords
Molecules, Peptides and proteins, Reaction mechanisms, Sulfides, Water
National Category
Organic Chemistry
Identifiers
urn:nbn:se:su:diva-215844 (URN)10.1021/acs.jpcb.2c08567 (DOI)000943621800001 ()36862530 (PubMedID)2-s2.0-85149506398 (Scopus ID)
Available from: 2023-03-29 Created: 2023-03-29 Last updated: 2023-03-29Bibliographically approved
Song, Y.-T., Li, X.-C. & Siegbahn, P. E. M. (2023). Is There a Different Mechanism for Water Oxidation in Higher Plants?. Journal of Physical Chemistry B, 127(30), 6643-6647
Open this publication in new window or tab >>Is There a Different Mechanism for Water Oxidation in Higher Plants?
2023 (English)In: Journal of Physical Chemistry B, ISSN 1520-6106, E-ISSN 1520-5207, Vol. 127, no 30, p. 6643-6647Article in journal (Refereed) Published
Abstract [en]

The leading mechanism for the formation of O-2 in photosystem II (PSII) has, during the past decade, been established as the so-called oxyl-oxo mechanism. In that mechanism, O-2 is formed from a binding between an oxygen radical (oxyl) and a bridging oxo group. For the case of higher plants, that mechanism has recently been criticized. Instead, a nucleophilic attack of an oxo group on a five-coordinated Mn(V)=O group forming O-2 has been suggested in a so-called water-unbound (WU) mechanism. In the present study, the WU mechanism has been investigated. It is found that the WU mechanism is just a variant of a previously suggested mechanism but with a reactant and a transition state that have much higher energies. The addition of a water molecule on the empty site of the Mn(V)=O center is very exergonic and leads back to the previously suggested oxyl-oxo mechanism.

National Category
Chemical Sciences
Identifiers
urn:nbn:se:su:diva-221318 (URN)10.1021/acs.jpcb.3c03029 (DOI)001032185800001 ()37467375 (PubMedID)2-s2.0-85166442276 (Scopus ID)
Available from: 2023-09-19 Created: 2023-09-19 Last updated: 2023-09-19Bibliographically approved
Siegbahn, P. E. M. (2023). The mechanism for N2 activation in the E4 – state of nitrogenase. Physical Chemistry, Chemical Physics - PCCP, 25(35), 23602-23613
Open this publication in new window or tab >>The mechanism for N2 activation in the E4 – state of nitrogenase
2023 (English)In: Physical Chemistry, Chemical Physics - PCCP, ISSN 1463-9076, E-ISSN 1463-9084, Vol. 25, no 35, p. 23602-23613Article in journal (Refereed) Published
Abstract [en]

Nitrogenases take nitrogen from the air and reduce it to ammonia. It has long been known that N2 becomes activated after four reductions in the catalytic cycle, in the E4 state. Several mechanisms for the activation have been suggested. In the present study a previous mechanism has been revised based on recent experimental findings. In the present mechanism N2H2 is formed in E4. As in the previously suggested mechanism, there are four initial reductions before catalysis (the A-states), after which a sulfide is released and the first state in catalysis (E0) is formed. In E4, N2 becomes bound and protonated in the Fe1, Fe2, Fe4 region, in which the hydrides have left two electrons. The rate-limiting step is the formation of N2H by a hydrogen atom transfer from Cys275 to N2 bound to Fe4, concerted with an additional electron transfer from the cofactor. The mechanism fulfills all requirements set by experiments. The activation of N2 is preceded by a formation of H2 from two hydrides, the carbide is kinetically hindered from being protonated, the E4 state is reversible. An important aspect is the presence of a water molecule in the Fe2, Fe6 region. The non-allowed formations of H2 from a hydride and a proton have been investigated and found to have higher barriers than the allowed formation of H2 from two hydrides.

National Category
Physical Chemistry
Identifiers
urn:nbn:se:su:diva-222256 (URN)10.1039/d3cp02851h (DOI)001066578800001 ()37622205 (PubMedID)2-s2.0-85170221824 (Scopus ID)
Available from: 2023-10-11 Created: 2023-10-11 Last updated: 2024-03-26Bibliographically approved
Pang, Y.-J., Li, X.-C., Siegbahn, P. E. M., Chen, G.-J. & Tan, H.-W. (2023). Theoretical Study of the Catalytic Mechanism of the Cu-Only Superoxide Dismutase. Journal of Physical Chemistry B, 127(21), 4800-4807
Open this publication in new window or tab >>Theoretical Study of the Catalytic Mechanism of the Cu-Only Superoxide Dismutase
Show others...
2023 (English)In: Journal of Physical Chemistry B, ISSN 1520-6106, E-ISSN 1520-5207, Vol. 127, no 21, p. 4800-4807Article in journal (Refereed) Published
Abstract [en]

The catalytic mechanisms for the wild-type and the mutated Cu-only superoxide dismutase were studied using the hybrid density functional B3LYP and a quantum chemical cluster approach. Optimal protonation states of the active site were examined for each stage of the catalytic cycle. For both the reductive and the oxidative half-reactions, the arrival of the substrate O-2(center dot-) was found to be accompanied by a chargecompensating H+ with exergonicities of -15.4 kcal center dot mol and -4.7 kcal center dot mol, respectively. The second-sphere Glu-110 and first-sphere His-93 were suggested to be the transient protonation site for the reductive and the oxidative half-reactions, respectively, which collaborates with the hydrogen bonding water chain to position the substrate near the redox-active copper center. For the reductive half-reaction, the rate-limiting step was found to be the inner-sphere electron transfer from the partially coordinated O-2(center dot-) to Cu-II with a barrier of 8.1 kcal center dot mol. The formed O-2 is released from the active site with an exergonicity of -14.9 kcal center dot mol. For the oxidative half-reaction, the inner-sphere electron transfer from CuI to the partially coordinated O-2(center dot-) was found to be accompanied by the proton transfer from the protonated His-93 and barrierless. The rate-limiting step was found to be the second proton transfer from the protonated Glu-110 to HO2 with a barrier of 7.3 kcal center dot mol. The barriers are reasonably consistent with experimental activities, and a proton-transfer rate-limiting step in the oxidative half-reaction could explain the experimentally observed pH-dependence. For the E110Q CuSOD, Asp-113 was suggested to be likely to serve as the transient protonation site in the reductive half-reaction. The rate-limiting barriers were found to be 8.0 and 8.6 kcal center dot mol, respectively, which could explain the slightly lower performance of E110X mutants. The results were found to be stable, with respect to the percentage of exact exchange in B3LYP.

National Category
Biochemistry and Molecular Biology
Identifiers
urn:nbn:se:su:diva-229899 (URN)10.1021/acs.jpcb.3c02175 (DOI)001016579700001 ()37196177 (PubMedID)
Available from: 2024-05-30 Created: 2024-05-30 Last updated: 2024-05-30Bibliographically approved
Wei, W.-J. & Siegbahn, P. E. M. (2022). A Mechanism for Nitrogenase Including Loss of a Sulfide. Chemistry - A European Journal, 28(12), Article ID e202103745.
Open this publication in new window or tab >>A Mechanism for Nitrogenase Including Loss of a Sulfide
2022 (English)In: Chemistry - A European Journal, ISSN 0947-6539, E-ISSN 1521-3765, Vol. 28, no 12, article id e202103745Article in journal (Refereed) Published
Abstract [en]

Nitrogenase is the only enzyme in nature that can fix N-2 from the air. The active cofactor of the leading form of this enzyme contains seven irons and one molybdenum connected by sulfide bridges. In several recent experimental studies, it has been suggested that the cofactor is very flexible, and might lose one of its sulfides during catalysis. In this study, the possible loss of a sulfide has been investigated by model calculations. In previous studies, we have shown that there should be four activation steps before catalysis starts, and this study is based on that finding. It was found here that, after the four reductions in the activation steps, a sulfide will become very loosely bound and can be released in a quite exergonic step with a low barrier. The binding of N-2 has no part in that release. In our previous studies, we suggested that the central carbide should be protonated three times after the four activation steps. With the new finding, there will instead be a loss of a sulfide, as the barrier for the loss is much lower than the ones for protonating the carbide. Still, it is suggested here that the carbide will be protonated anyway, but only with one proton, in the E-3 to E-4 step. A very complicated transition state for H-2 formation involving a large structural change was obtained. The combined step, with a loss of H-2 and binding of N-2, is calculated to be endergonic by +2.3 kcal mol(-1); this is in excellent agreement with experiments in which an easily reversible step has been found.

Keywords
activation, density functional calculations, mechanisms, nitrogenases, potential surfaces, sulfides
National Category
Chemical Sciences
Identifiers
urn:nbn:se:su:diva-202394 (URN)10.1002/chem.202103745 (DOI)000750229300001 ()35098591 (PubMedID)
Available from: 2022-03-03 Created: 2022-03-03 Last updated: 2022-03-03Bibliographically approved
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