Inhibitor Binding Modulates Protonation States in the Active Site of SARS-CoV-2 Main Protease

2021 ◽  
Author(s):  
Daniel W. Kneller ◽  
Gwyndalyn Phillips ◽  
Kevin L. Weiss ◽  
Qiu Zhang ◽  
Leighton Coates ◽  
...  
Keyword(s):  
Active Site ◽  
Rational Drug Design ◽  
Inhibitor Binding ◽  
Histidine Residues ◽  
Main Protease ◽  
Rational Drug ◽  
Ligand Free ◽  
Essential Enzyme ◽  
Neutron Protein Crystallography ◽  
Active Site Histidine

ABSTRACTThe main protease (3CL Mpro) from SARS-CoV-2, the virus that causes COVID-19, is an essential enzyme for viral replication with no human counterpart, making it an attractive drug target. Although drugs have been developed to inhibit the proteases from HIV, hepatitis C and other viruses, no such therapeutic is available to inhibit the main protease of SARS-CoV-2. To directly observe the protonation states in SARS-CoV-2 Mpro and to elucidate their importance in inhibitor binding, we determined the structure of the enzyme in complex with the α-ketoamide inhibitor telaprevir using neutron protein crystallography at near-physiological temperature. We compared protonation states in the inhibitor complex with those determined for a ligand-free neutron structure of Mpro. This comparison revealed that three active-site histidine residues (His41, His163 and His164) adapt to ligand binding, altering their protonation states to accommodate binding of telaprevir. We suggest that binding of other α-ketoamide inhibitors can lead to the same protonation state changes of the active site histidine residues. Thus, by studying the role of active site protonation changes induced by inhibitors we provide crucial insights to help guide rational drug design, allowing precise tailoring of inhibitors to manipulate the electrostatic environment of SARS-CoV-2 Mpro.

scholarly journals Protonation states in SARS-CoV-2 main protease mapped by neutron crystallography

2020 ◽  
Author(s):  
Daniel W. Kneller ◽  
Gwyndalyn Phillips ◽  
Kevin L. Weiss ◽  
Swati Pant ◽  
Qiu Zhang ◽  
...  
Keyword(s):  
Active Site ◽  
Direct Determination ◽  
Amino Acid Residues ◽  
Main Protease ◽  
Neutron Structure ◽  
Active Site Cavity ◽  
Ionizable Residues ◽  
Essential Enzyme ◽  
Neutron Protein Crystallography

AbstractThe main protease (3CL Mpro) from SARS-CoV-2, the etiological agent of COVID-19, is an essential enzyme for viral replication, possessing an unusual catalytic dyad composed of His41 and Cys145. A long-standing question in the field has been what the protonation states of the ionizable residues in the substrate-binding active site cavity are. Here, we present the room-temperature neutron structure of 3CL Mpro from SARS-CoV-2, which allows direct determination of hydrogen atom positions and, hence, protonation states. The catalytic site natively adopts a zwitterionic reactive state where His41 is doubly protonated and positively charged, and Cys145 is in the negatively charged thiolate state. The neutron structure also identified the protonation states of other amino acid residues, mapping electrical charges and intricate hydrogen bonding networks in the SARS-CoV-2 3CL Mpro active site cavity and dimer interface. This structure highlights the ability of neutron protein crystallography for experimentally determining protonation states at near-physiological temperature – the critical information for structure-assisted and computational drug design.

scholarly journals The dapE-encoded N-succinyl-l,l-diaminopimelic acid desuccinylase from Haemophilus influenzae contains two active-site histidine residues

2008 ◽  
Vol 14 (1) ◽  
pp. 1-10 ◽  
Author(s):  
Danuta M. Gillner ◽  
David L. Bienvenue ◽  
Boguslaw P. Nocek ◽  
Andrzej Joachimiak ◽  
Vincentos Zachary ◽  
...  
Keyword(s):  
Haemophilus Influenzae ◽  
Active Site ◽  
Diaminopimelic Acid ◽  
Histidine Residues ◽  
Active Site Histidine

scholarly journals Estudo QM/MM da proteína tirosina Bcr-Abl mutada T315I

2020 ◽  
Author(s):  
Washington A. Pereira ◽  
Érica C. M. Nascimento ◽  
João B. L. Martins
Keyword(s):  
Active Site ◽  
Myeloid Leukemia ◽  
Empirical Method ◽  
Rational Drug Design ◽  
Signaling Cascade ◽  
Amber Force Field ◽  
Abl Tyrosine Kinase ◽  
Rational Drug ◽  
High Layer ◽  
Semi Empirical

Bcr-Abl tyrosine kinase protein activates the substrate starting kinase signaling cascade that results in cell division. When in excess this activation can lead to chronic myeloid leukemia. Currently, the treatment of this disease is achieved through drugs developed by rational drug design, e.g., imatinib. In this work we study descriptors of the following molecules: imatinib, nilotinib and ponatinib, together with a mutated protein. To characterize interactions between the residues of the active site against those inhibitors, a QM/MM study was carried out, using the hybrid method ONIOM, through the AMBER Force field (low layer) and the semi-empirical method PM6 (high layer).

scholarly journals IN Silico Approach of Some Selected Honey Constituents as SARS-CoV-2 Main Protease (COVID-19) Inhibitors

2020 ◽  
Author(s):  
Heba Hashem
Keyword(s):  
Active Site ◽  
Life Cycles ◽  
High Binding Energy ◽  
Protease Inhibition ◽  
Enzyme Active Site ◽  
Therapeutic Drugs ◽  
Main Protease ◽  
The World ◽  
Essential Enzyme ◽  
The Way

<p>The huge attack of coronavirus disease 2019 (COVID-19) over all the world forces the researcher around the world to study the crystal structure of the main protease M<sup>pro</sup> ( 3-chymotrypsin-like cysteine enzyme) which is the essential enzyme for coronavirus processing the polyproteins and its life cycles. And by the way, the inhibition of this enzyme active site becomes the target of all scientists of drug discovery in order to overcome this disease. In this study, we have used the molecular modeling approach to evaluate the activity of different active compounds from honeybee and propolis to inhibit the presented sars-cov-2 main protease via Schrödinger Maestro v10.1. the presented study resulted in six main compounds possess high binding energy with the receptor active site of COVID-19 main protease. we hope this study being the way for honeybee constitution as an effective ligand for sars-cov-2 main protease inhibition and be in the medicinal study of anti-COVID-19 therapeutic drugs.</p>

scholarly journals Identification of Active-Site Histidine Residues of a Self-Incompatibility Ribonuclease from a Wild Tomato

1997 ◽  
Vol 115 (4) ◽  
pp. 1421-1429 ◽  
Author(s):  
S. Parry ◽  
E. Newbigin ◽  
G. Currie ◽  
A. Bacic ◽  
D. Oxley
Keyword(s):  
Active Site ◽  
Self Incompatibility ◽  
Histidine Residues ◽  
Wild Tomato ◽  
Active Site Histidine

scholarly journals Phenothiazines as dual inhibitors of SARS-CoV-2 main protease and COVID-19 inflammation

2021 ◽  
Author(s):  
Katrina L Forrestall ◽  
Darcy E Burley ◽  
Meghan Kirsten Cash ◽  
Ian Pottie ◽  
Sultan Darvesh
Keyword(s):  
Viral Replication ◽  
Active Site ◽  
Acute Infection ◽  
Binding Energies ◽  
Neurotransmitter Receptors ◽  
Main Protease ◽  
Dual Action ◽  
Inhibition Constants ◽  
Anti Inflammatory ◽  
Essential Enzyme

COVID-19, caused by the severe acute respiratory coronavirus 2 (SARS-CoV-2) currently has no treatment for acute infection. The main protease (Mpro) of SARS-CoV-2 is an essential enzyme for viral replication and an attractive target for disease intervention. The phenothiazine moiety has demonstrated drug versatility for biological systems, including inhibition of butyrylcholinesterase, a property important in the cholinesterase anti-inflammatory cascade. Nineteen phenothiazine drugs were investigated using in silico modelling techniques to predict binding energies and inhibition constants (Ki values) with SARS-CoV-2 Mpro. Since most side-effects of phenothiazines are due to interactions with various neurotransmitter receptors and transporters, phenothiazines with few such interactions were also investigated. All compounds were found to bind to the active site of SARS-CoV-2 Mpro and showed Ki values ranging from 1.30 to 52.4 µM. Nine phenothiazines showed inhibition constants <10 µM. The compounds with limited interactions with neurotransmitter receptors and transporters showed micromolar (µM) Ki values. Docking results were compared with remdesivir and showed similar interactions with key residues Glu-166 and Gln-189 in the active site. This work has identified several phenothiazines with limited neurotransmitter receptor and transporter interactions and that may provide the dual action of inhibiting SARS-CoV-2 Mpro to prevent viral replication and promote the release of anti-inflammatory cytokines to curb viral-induced inflammation. These compounds are promising candidates for further investigation against SARS-CoV-2.

scholarly journals Unusual zwitterionic catalytic site of SARS–CoV-2 main protease revealed by neutron crystallography

2020 ◽  
Vol 295 (50) ◽  
pp. 17365-17373 ◽  
Author(s):  
Daniel W. Kneller ◽  
Gwyndalyn Phillips ◽  
Kevin L. Weiss ◽  
Swati Pant ◽  
Qiu Zhang ◽  
...  
Keyword(s):  
Active Site ◽  
Direct Determination ◽  
Catalytic Site ◽  
Amino Acid Residues ◽  
Critical Question ◽  
Main Protease ◽  
Neutron Structure ◽  
Active Site Cavity ◽  
Ionizable Residues ◽  
Essential Enzyme

The main protease (3CL Mpro) from SARS–CoV-2, the etiological agent of COVID-19, is an essential enzyme for viral replication. 3CL Mpro possesses an unusual catalytic dyad composed of Cys145 and His41 residues. A critical question in the field has been what the protonation states of the ionizable residues in the substrate-binding active-site cavity are; resolving this point would help understand the catalytic details of the enzyme and inform rational drug development against this pernicious virus. Here, we present the room-temperature neutron structure of 3CL Mpro, which allowed direct determination of hydrogen atom positions and, hence, protonation states in the protease. We observe that the catalytic site natively adopts a zwitterionic reactive form in which Cys145 is in the negatively charged thiolate state and His41 is doubly protonated and positively charged, instead of the neutral unreactive state usually envisaged. The neutron structure also identified the protonation states, and thus electrical charges, of all other amino acid residues and revealed intricate hydrogen-bonding networks in the active-site cavity and at the dimer interface. The fine atomic details present in this structure were made possible by the unique scattering properties of the neutron, which is an ideal probe for locating hydrogen positions and experimentally determining protonation states at near-physiological temperature. Our observations provide critical information for structure-assisted and computational drug design, allowing precise tailoring of inhibitors to the enzyme's electrostatic environment.

scholarly journals Malate dehydrogenase of the cytosol. Ionizations of the enzyme-reduced-coenzyme complex and a comparison with lactate dehydrogenase

1978 ◽  
Vol 173 (2) ◽  
pp. 597-605 ◽  
Author(s):  
A Lodola ◽  
D M Parker ◽  
R Jeck ◽  
J J Holbrook
Keyword(s):  
Lactate Dehydrogenase ◽  
Malate Dehydrogenase ◽  
Active Site ◽  
Histidine Residue ◽  
Histidine Residues ◽  
Coenzyme Binding ◽  
Active Site Histidine

1. The pH-dependencies of the binding of NADH and reduced nicotinamide–benzimidazole dinucleotide to pig heart cytoplasmic malate dehydrogenase and lactate dehydrogenase are reported. 2. Two ionizing groups were observed in the binding of both reduced coenzymes to lactate dehydrogenase. One group, with pKa in the range 6.3–6.7, is the active-site histidine residue and its deprotonation weakens binding of reduced coenzyme 3-fold. Binding of both coenzymes is decreased to zero when a second group, of pKa 8.9, deprotonates. This group is not cysteine-165.3. Only one ionization is required to characterize the binding of the two reduced coenzymes to malate dehydrogenase. The group involved appears to be the active-site histidine residue, since its ethoxycarbonylation inhibits the enzyme and abolishes binding of reduced coenzyme. Binding of either reduced coenzyme increases the pKa of the group from 6.4 to 7.4, and deprotonation of the group is accompanied by a 10-fold weakening of coenzyme binding. 4. Two reactive histidine residues were detected per malate dehydrogenase dimer. 5. A mechanism which emphasizes the homology between the two enzymes is presented.

scholarly journals Crystal structure correlations with the intrinsic thermodynamics of human carbonic anhydrase inhibitor binding

PeerJ ◽  
2018 ◽  
Vol 6 ◽  
pp. e4412 ◽  
Author(s):  
Alexey Smirnov ◽  
Asta Zubrienė ◽  
Elena Manakova ◽  
Saulius Gražulis ◽  
Daumantas Matulis
Keyword(s):  
Carbonic Anhydrase ◽  
Structural Information ◽  
Hydrophobic Effect ◽  
Rational Drug Design ◽  
Inhibitor Binding ◽  
Binding Thermodynamics ◽  
Chemical Structures ◽  
Structure Correlations ◽  
Rational Drug ◽  
Human Carbonic Anhydrase

The structure-thermodynamics correlation analysis was performed for a series of fluorine- and chlorine-substituted benzenesulfonamide inhibitors binding to several human carbonic anhydrase (CA) isoforms. The total of 24 crystal structures of 16 inhibitors bound to isoforms CA I, CA II, CA XII, and CA XIII provided the structural information of selective recognition between a compound and CA isoform. The binding thermodynamics of all structures was determined by the analysis of binding-linked protonation events, yielding the intrinsic parameters, i.e., the enthalpy, entropy, and Gibbs energy of binding. Inhibitor binding was compared within structurally similar pairs that differ bypara-ormeta-substituents enabling to obtain the contributing energies of ligand fragments. The pairs were divided into two groups. First,similarbinders—the pairs that keep the same orientation of the benzene ring exhibited classical hydrophobic effect, a less exothermic enthalpy and a more favorable entropy upon addition of the hydrophobic fragments. Second,dissimilarbinders—the pairs of binders that demonstrated altered positions of the benzene rings exhibited the non-classical hydrophobic effect, a more favorable enthalpy and variable entropy contribution. A deeper understanding of the energies contributing to the protein-ligand recognition should lead toward the eventual goal of rational drug design where chemical structures of ligands could be designed based on the target protein structure.

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