Quantum-Chemical Study of the Discrimination against dNTP in the Nucleotide Addition Reaction in the Active Site of RNA Polymerase II

2017 ◽  
Vol 13 (4) ◽  
pp. 1699-1705 ◽  
Author(s):  
Sven Roßbach ◽  
Christian Ochsenfeld
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Active Site ◽  
Quantum Chemical ◽  
Addition Reaction ◽  
Quantum Chemical Study ◽  
Chemical Study ◽  
Nucleotide Addition

scholarly journals Structures of E. coli σS-transcription initiation complexes provide new insights into polymerase mechanism

2016 ◽  
Vol 113 (15) ◽  
pp. 4051-4056 ◽  
Author(s):  
Bin Liu ◽  
Yuhong Zuo ◽  
Thomas A. Steitz
Keyword(s):  
Rna Polymerase ◽  
Conformational Changes ◽  
Active Site ◽  
Transcription Initiation ◽  
De Novo ◽  
Addition Reaction ◽  
Specific Interactions ◽  
Promoter Recognition ◽  
Nucleotide Addition ◽  
Promoter Dna

In bacteria, multiple σ factors compete to associate with the RNA polymerase (RNAP) core enzyme to form a holoenzyme that is required for promoter recognition. During transcription initiation RNAP remains associated with the upstream promoter DNA via sequence-specific interactions between the σ factor and the promoter DNA while moving downstream for RNA synthesis. As RNA polymerase repetitively adds nucleotides to the 3′-end of the RNA, a pyrophosphate ion is generated after each nucleotide incorporation. It is currently unknown how the release of pyrophosphate affects transcription. Here we report the crystal structures of E. coli transcription initiation complexes (TICs) containing the stress-responsive σS factor, a de novo synthesized RNA oligonucleotide, and a complete transcription bubble (σS-TIC) at about 3.9-Å resolution. The structures show the 3D topology of the σS factor and how it recognizes the promoter DNA, including likely specific interactions with the template-strand residues of the −10 element. In addition, σS-TIC structures display a highly stressed pretranslocated initiation complex that traps a pyrophosphate at the active site that remains closed. The position of the pyrophosphate and the unusual phosphodiester linkage between the two terminal RNA residues suggest an unfinished nucleotide-addition reaction that is likely at equilibrium between nucleotide addition and pyrophosphorolysis. Although these σS-TIC crystals are enzymatically active, they are slow in nucleotide addition, as suggested by an NTP soaking experiment. Pyrophosphate release completes the nucleotide addition reaction and is associated with extensive conformational changes around the secondary channel but causes neither active site opening nor transcript translocation.

A Quantum Chemical Study of the Synthesis of Prostaglandin G2by the Cyclooxygenase Active Site in Prostaglandin Endoperoxide H Synthase 1

2003 ◽  
Vol 107 (14) ◽  
pp. 3297-3308 ◽  
Author(s):  
L. Mattias Blomberg ◽  
Margareta R. A. Blomberg ◽  
Per E. M. Siegbahn ◽  
Wilfred A. van der Donk ◽  
Ah-Lim Tsai
Keyword(s):  
Active Site ◽  
Quantum Chemical ◽  
Quantum Chemical Study ◽  
Chemical Study ◽  
Prostaglandin Endoperoxide ◽  
Endoperoxide H Synthase

Quantum-chemical study of the photochemical addition reaction of dialkyl disulfides with olefins

2009 ◽  
Vol 49 (3) ◽  
pp. 240-244
Author(s):  
Yu. A. Borisov ◽  
A. K. Dyusengaliev ◽  
G. A. Orazova ◽  
T. P. Serikov
Keyword(s):  
Quantum Chemical ◽  
Addition Reaction ◽  
Quantum Chemical Study ◽  
Chemical Study ◽  
Dialkyl Disulfides

scholarly journals Nucleotide Loading Modes of Human RNA Polymerase II as Deciphered by Molecular Simulations

Biomolecules ◽  
2020 ◽  
Vol 10 (9) ◽  
pp. 1289
Author(s):  
Nicolas E. J. Génin ◽  
Robert O. J. Weinzierl
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Active Site ◽  
Nucleoside Triphosphate ◽  
Accelerated Molecular Dynamics ◽  
Structural Framework ◽  
Catalytic Isomerization ◽  
Nucleotide Addition ◽  
Loading Modes ◽  
Dynamics Simulations

Mapping the route of nucleoside triphosphate (NTP) entry into the sequestered active site of RNA polymerase (RNAP) has major implications for elucidating the complete nucleotide addition cycle. Constituting a dichotomy that remains to be resolved, two alternatives, direct NTP delivery via the secondary channel (CH2) or selection to downstream sites in the main channel (CH1) prior to catalysis, have been proposed. In this study, accelerated molecular dynamics simulations of freely diffusing NTPs about RNAPII were applied to refine the CH2 model and uncover atomic details on the CH1 model that previously lacked a persuasive structural framework to illustrate its mechanism of action. Diffusion and binding of NTPs to downstream DNA, and the transfer of a preselected NTP to the active site, are simulated for the first time. All-atom simulations further support that CH1 loading is transcription factor IIF (TFIIF) dependent and impacts catalytic isomerization. Altogether, the alternative nucleotide loading systems may allow distinct transcriptional landscapes to be expressed.

scholarly journals Hydroxide Once or Twice? A combined Neutron Crystallographic and Quantum Chemical Study of the Hydride Shift in D-Xylose Isomerase

2015 ◽  
Author(s):  
Matt Challacombe ◽  
Nicolas Bock ◽  
Paul Langan ◽  
Andrey Kovalevsky
Keyword(s):  
Branch Point ◽  
Active Site ◽  
Quantum Chemical ◽  
Large Scale ◽  
Quantum Chemical Study ◽  
Xylose Isomerase ◽  
Chemical Study ◽  
Rigid Boundary ◽  
Hydride Shift ◽  
Additional Hydrogen Bond

The hydride shift mechanism of D-Xylose Isomerase converts D-glucose to D-fructose. In this work, we compute features of a “hydroxide once” mechanism for the hydride shift with quantum chemical calculations based on the 3KCO (linear) and 3KCL (cyclic) X-ray/neutron structures. The rigid boundary conditions of the active site “shoe-box”, together with ionization states and proton orientations, enables large scale electronic structure calculations of the entire active site with greatly reduced configuration sampling. In the reported hydroxide once mechanism, magnesium in the 2A ligation shifts to position 2B, ionizing the O2 proton of D-glucose, which is accepted by ASP-287. In this step a novel stabilization is discovered; the K183/D255 proton toggle, providing a ∼10 kcal/mol stabilization through inductive polarization over 5Å. Then, hydride shifts from glucose-O2 to glucose-O1 (the interconversion) generating hydroxide (once) from the catalytic water. This step is consistent with the observation of hyroxide in structure 3CWH, which we identify as a branch point. From this branch point, we find several routes to the solvent-free regeneration of catalytic water that is strongly exothermic (by ∼20 kcal/mol), yielding one additional hydrogen bond more than the starting structure. This non-Michaelis behavior, strongly below the starting cyclic and linear total energy, explains the observed accumulation of hydroxide intermediate -- we postulate that forming permissive ionization states, required for cyclization, may be the rate limiting step. In all, we find eight items of experimental correspondence supporting features of the putative hydroxide once mechanism.

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