Oncogenic G12D mutation alters local conformations and dynamics of K-Ras

AbstractK-Ras is the most frequently mutated oncoprotein in human cancers, and G12D is its most prevalent mutation. To understand how G12D mutation impacts K-Ras function, we need to understand how it alters the regulation of its dynamics. Here, we present local changes in K-Ras structure, conformation and dynamics upon G12D mutation, from long-timescale Molecular Dynamics simulations of active (GTP-bound) and inactive (GDP-bound) forms of wild-type and mutant K-Ras, with an integrated investigation of atomistic-level changes, local conformational shifts and correlated residue motions. Our results reveal that the local changes in K-Ras are specific to bound nucleotide (GTP or GDP), and we provide a structural basis for this. Specifically, we show that G12D mutation causes a shift in the population of local conformational states of K-Ras, especially in Switch-II (SII) and α3-helix regions, in favor of a conformation that is associated with a catalytically impaired state through structural changes; it also causes SII motions to anti-correlate with other regions. This detailed picture of G12D mutation effects on the local dynamic characteristics of both active and inactive protein helps enhance our understanding of local K-Ras dynamics, and can inform studies on the development of direct inhibitors towards the treatment of K-RasG12D-driven cancers.

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Discovery of natural product ovalicin sensitive type 1 methionine aminopeptidases: molecular and structural basis

Biochemical Journal ◽

10.1042/bcj20180874 ◽

2019 ◽

Vol 476 (6) ◽

pp. 991-1003 ◽

Cited By ~ 1

Author(s):

Vijaykumar Pillalamarri ◽

Tarun Arya ◽

Neshatul Haque ◽

Sandeep Chowdary Bala ◽

Anil Kumar Marapaka ◽

...

Keyword(s):

Natural Product ◽

Active Site ◽

Covalent Modification ◽

Structural Basis ◽

Wild Type ◽

Methionine Aminopeptidases ◽

Synthetic Derivative ◽

Dynamics Simulations

Abstract Natural product ovalicin and its synthetic derivative TNP-470 have been extensively studied for their antiangiogenic property, and the later reached phase 3 clinical trials. They covalently modify the conserved histidine in Type 2 methionine aminopeptidases (MetAPs) at nanomolar concentrations. Even though a similar mechanism is possible in Type 1 human MetAP, it is inhibited only at millimolar concentration. In this study, we have discovered two Type 1 wild-type MetAPs (Streptococcus pneumoniae and Enterococcus faecalis) that are inhibited at low micromolar to nanomolar concentrations and established the molecular mechanism. F309 in the active site of Type 1 human MetAP (HsMetAP1b) seems to be the key to the resistance, while newly identified ovalicin sensitive Type 1 MetAPs have a methionine or isoleucine at this position. Type 2 human MetAP (HsMetAP2) also has isoleucine (I338) in the analogous position. Ovalicin inhibited F309M and F309I mutants of human MetAP1b at low micromolar concentration. Molecular dynamics simulations suggest that ovalicin is not stably placed in the active site of wild-type MetAP1b before the covalent modification. In the case of F309M mutant and human Type 2 MetAP, molecule spends more time in the active site providing time for covalent modification.

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Mechanistic Insights into the Effects of Key Mutations on SARS-CoV-2 RBD-ACE2 Binding

10.1101/2021.09.20.461041 ◽

2021 ◽

Author(s):

Abhishek Aggarwal ◽

Supriyo Naskar ◽

Nikhil Maroli ◽

Biswajit Gorai ◽

Narendra M Dixit ◽

...

Keyword(s):

Structural Changes ◽

Underlying Mechanism ◽

Free Energy Calculations ◽

Target Cells ◽

Original Strain ◽

Binding Affinities ◽

Receptor Binding Domain ◽

Wild Type ◽

Dynamics Simulations ◽

Insight Into

Some recent SARS-CoV-2 variants appear to have increased transmissibility than the original strain. An underlying mechanism could be the improved ability of the variants to bind receptors on target cells and infect them. In this study, we provide atomic-level insight into the binding of the receptor binding domain (RBD) of the wild-type SARS-CoV-2 spike protein and its single (N501Y), double (E484Q, L452R) and triple (N501Y, E484Q, L452R) mutated variants to the human ACE2 receptor. Using extensive all-atom molecular dynamics simulations and advanced free energy calculations, we estimate the associated binding affinities and binding hotspots. We observe significant secondary structural changes in the RBD of the mutants, which lead to different binding affinities. We find higher binding affinities of the double (E484Q, L452R) and triple (N501Y, E484Q, L452R) mutated variants than the wild type and the N501Y variant, which could contribute to the higher transmissibility of recent variants containing these mutations.

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Structural basis of the regulation of the normal and oncogenic methylation of nucleosomal histone H3 Lys36 by NSD2

Nature Communications ◽

10.1038/s41467-021-26913-5 ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Ko Sato ◽

Amarjeet Kumar ◽

Keisuke Hamada ◽

Chikako Okada ◽

Asako Oguni ◽

...

Keyword(s):

Histone H3 ◽

Structural Basis ◽

Wild Type ◽

Cryo Electron Microscopy ◽

Nucleosomal Dna ◽

Oncogenic Mutations ◽

Binding Cleft ◽

Dynamics Simulations ◽

Nucleosome Binding ◽

Nucleosomal Histone

AbstractDimethylated histone H3 Lys36 (H3K36me2) regulates gene expression, and aberrant H3K36me2 upregulation, resulting from either the overexpression or point mutation of the dimethyltransferase NSD2, is found in various cancers. Here we report the cryo-electron microscopy structure of NSD2 bound to the nucleosome. Nucleosomal DNA is partially unwrapped, facilitating NSD2 access to H3K36. NSD2 interacts with DNA and H2A along with H3. The NSD2 autoinhibitory loop changes its conformation upon nucleosome binding to accommodate H3 in its substrate-binding cleft. Kinetic analysis revealed that two oncogenic mutations, E1099K and T1150A, increase NSD2 catalytic turnover. Molecular dynamics simulations suggested that in both mutants, the autoinhibitory loop adopts an open state that can accommodate H3 more often than the wild-type. We propose that E1099K and T1150A destabilize the interactions that keep the autoinhibitory loop closed, thereby enhancing catalytic turnover. Our analyses guide the development of specific inhibitors of NSD2.

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Cryo-EM structures of undocked innexin-6 hemichannels in phospholipids

Science Advances ◽

10.1126/sciadv.aax3157 ◽

2020 ◽

Vol 6 (7) ◽

pp. eaax3157 ◽

Cited By ~ 3

Author(s):

Batuujin Burendei ◽

Ruriko Shinozaki ◽

Masakatsu Watanabe ◽

Tohru Terada ◽

Kazutoshi Tani ◽

...

Keyword(s):

Gap Junctions ◽

Structural Basis ◽

Wild Type ◽

Amino Terminal ◽

Functional Assays ◽

Cryo Electron Microscopy ◽

Dynamics Simulations ◽

Open States ◽

Insight Into ◽

Junction Proteins

Gap junctions form intercellular conduits with a large pore size whose closed and open states regulate communication between adjacent cells. The structural basis of the mechanism by which gap junctions close, however, remains uncertain. Here, we show the cryo–electron microscopy structures of Caenorhabditis elegans innexin-6 (INX-6) gap junction proteins in an undocked hemichannel form. In the nanodisc-reconstituted structure of the wild-type INX-6 hemichannel, flat double-layer densities obstruct the channel pore. Comparison of the hemichannel structures of a wild-type INX-6 in detergent and nanodisc-reconstituted amino-terminal deletion mutant reveals that lipid-mediated amino-terminal rearrangement and pore obstruction occur upon nanodisc reconstitution. Together with molecular dynamics simulations and electrophysiology functional assays, our results provide insight into the closure of the INX-6 hemichannel in a lipid bilayer before docking of two hemichannels.

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Structural Basis for Regioselectivity and Stereoselectivity of Product Formation by Naphthalene 1,2-Dioxygenase

Journal of Bacteriology ◽

10.1128/jb.00707-06 ◽

2006 ◽

Vol 188 (19) ◽

pp. 6986-6994 ◽

Cited By ~ 35

Author(s):

Daniel J. Ferraro ◽

Adam L. Okerlund ◽

Jonathan C. Mowers ◽

S. Ramaswamy

Keyword(s):

Active Site ◽

Structural Changes ◽

Product Formation ◽

Structural Basis ◽

Wild Type ◽

Native Form ◽

Rational Engineering ◽

Aromatic Substrates ◽

Primary Determinant ◽

In The Wild

ABSTRACT Rieske oxygenase (RO) systems are two- and three-component enzyme systems that catalyze the formation of cis-dihydrodiols from aromatic substrates. Degradation of pollutants in contaminated soil and generation of chiral synthons have been the major foci of RO research. Substrate specificity and product regio- and stereoselectivity have been shown to vary between individual ROs. While directed evolution methods for altering RO function have been successful in the past, rational engineering of these enzymes still poses a challenge due to the lack of structural understanding. Here we examine the structural changes induced by mutation of Phe-352 in naphthalene 1,2-dioxygenase from Pseudomonas sp. strain NCIB 9816-4 (NDO-O9816-4). Structures of the Phe-352-Val mutant in native form and in complex with phenanthrene and anthracene, along with those of wild-type NDO-O9816-4 in complex with phenanthrene, anthracene, and 3-nitrotoluene, are presented. Phenanthrene was shown to bind in a different orientation in the Phe-352-Val mutant active site from that in the wild type, while anthracene was found to bind in similar positions in both enzymes. Two orientations of 3-nitrotoluene were observed, i.e., a productive and a nonproductive orientation. These orientations help explain why NDO-O9816-4 forms different products from 3-nitrotoluene than those made from nitrobenzene dioxygenase. Comparison of these structures among themselves and with other known ROs bound to substrates reveals that the orientation of substrate binding at the active site is the primary determinant of product regio- and stereoselectivity.

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Structural basis for the reaction cycle of DASS dicarboxylate transporters

eLife ◽

10.7554/elife.61350 ◽

2020 ◽

Vol 9 ◽

Cited By ~ 2

Author(s):

David B Sauer ◽

Noah Trebesch ◽

Jennifer J Marden ◽

Nicolette Cocco ◽

Jinmei Song ◽

...

Keyword(s):

Structural Changes ◽

Tca Cycle ◽

Charge Compensation ◽

Structural Basis ◽

Cellular Regulation ◽

Transport Cycle ◽

Tca Cycle Intermediates ◽

Dynamics Simulations ◽

A Charge ◽

Transport Domain

Citrate, α-ketoglutarate and succinate are TCA cycle intermediates that also play essential roles in metabolic signaling and cellular regulation. These di- and tricarboxylates are imported into the cell by the divalent anion sodium symporter (DASS) family of plasma membrane transporters, which contains both cotransporters and exchangers. While DASS proteins transport substrates via an elevator mechanism, to date structures are only available for a single DASS cotransporter protein in a substrate-bound, inward-facing state. We report multiple cryo-EM and X-ray structures in four different states, including three hitherto unseen states, along with molecular dynamics simulations, of both a cotransporter and an exchanger. Comparison of these outward- and inward-facing structures reveal how the transport domain translates and rotates within the framework of the scaffold domain through the transport cycle. Additionally, we propose that DASS transporters ensure substrate coupling by a charge-compensation mechanism, and by structural changes upon substrate release.

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Biochemical and Structural Characterization of OXA-405, an OXA-48 Variant with Extended-Spectrum β-Lactamase Activity

Microorganisms ◽

10.3390/microorganisms8010024 ◽

2019 ◽

Vol 8 (1) ◽

pp. 24 ◽

Cited By ~ 4

Author(s):

Saoussen Oueslati ◽

Pascal Retailleau ◽

Ludovic Marchini ◽

Laurent Dortet ◽

Rémy A. Bonnin ◽

...

Keyword(s):

Substrate Specificity ◽

Structural Changes ◽

3D Structure ◽

Hydrolytic Activity ◽

Structural Basis ◽

Class D ◽

Covalent Docking ◽

Dynamics Simulations ◽

Hydrolysis Of

OXA-48-producing Enterobacterales have now widely disseminated globally. A sign of their extensive spread is the identification of an increasing number of OXA-48 variants. Among them, three are particularly interesting, OXA-163, OXA-247 and OXA-405, since they have lost carbapenem activities and gained expanded-spectrum cephalosporin hydrolytic activity subsequent to a four amino-acid (AA) deletion in the β5–β6 loop. We investigated the mechanisms responsible for substrate specificity of OXA-405. Kinetic parameters confirmed that OXA-405 has a hydrolytic profile compatible with an ESBL (hydrolysis of expanded spectrum cephalosporins and susceptibility to class A inhibitors). Molecular modeling techniques and 3D structure determination show that the overall dimeric structure of OXA-405 is very similar to that of OXA-48, except for the β5–β6 loop, which is shorter for OXA-405, suggesting that the length of the β5–β6 loop is critical for substrate specificity. Covalent docking with selected substrates and molecular dynamics simulations evidenced the structural changes induced by substrate binding, as well as the distribution of water molecules in the active site and their role in substrate hydrolysis. All this data may represent the structural basis for the design of new and efficient class D inhibitors.

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Altered Local Interactions and Long-Range Communications in UK Variant (B.1.1.7) Spike Glycoprotein

International Journal of Molecular Sciences ◽

10.3390/ijms22115464 ◽

2021 ◽

Vol 22 (11) ◽

pp. 5464

Author(s):

Stefano Borocci ◽

Carmen Cerchia ◽

Alessandro Grottesi ◽

Nico Sanna ◽

Ingrid Guarnetti Prandi ◽

...

Keyword(s):

Neutralizing Antibodies ◽

Structural Information ◽

Conformational Transitions ◽

Structural Basis ◽

Receptor Binding Domain ◽

Wild Type ◽

Dynamics Simulations ◽

The Uk ◽

The Impact

The COVID-19 pandemic is caused by SARS-CoV-2. Currently, most of the research efforts towards the development of vaccines and antibodies against SARS-CoV-2 were mainly focused on the spike (S) protein, which mediates virus entry into the host cell by binding to ACE2. As the virus SARS-CoV-2 continues to spread globally, variants have emerged, characterized by multiple mutations of the S glycoprotein. Herein, we employed microsecond-long molecular dynamics simulations to study the impact of the mutations of the S glycoprotein in SARS-CoV-2 Variant of Concern 202012/01 (B.1.1.7), termed the “UK variant”, in comparison with the wild type, with the aim to decipher the structural basis of the reported increased infectivity and virulence. The simulations provided insights on the different dynamics of UK and wild-type S glycoprotein, regarding in particular the Receptor Binding Domain (RBD). In addition, we investigated the role of glycans in modulating the conformational transitions of the RBD. The overall results showed that the UK mutant experiences higher flexibility in the RBD with respect to wild type; this behavior might be correlated with the increased transmission reported for this variant. Our work also adds useful structural information on antigenic “hotspots” and epitopes targeted by neutralizing antibodies.

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Tuning the Electric Field Response of MOFs by Rotatable Dipolar Linkers

10.26434/chemrxiv.8153198.v1 ◽

2019 ◽

Author(s):

Johannes P. Dürholt ◽

Babak Farhadi Jahromi ◽

Rochus Schmid

Keyword(s):

Electric Field ◽

Electric Fields ◽

Structural Changes ◽

Dielectric Behavior ◽

Two Dimensions ◽

Dielectric Constants ◽

Electric Response ◽

Dipole Strength ◽

Metal Organic ◽

Dynamics Simulations

Recently the possibility of using electric fields as a further stimulus to trigger structural changes in metal-organic frameworks (MOFs) has been investigated. In general, rotatable groups or other types of mechanical motion can be driven by electric fields. In this study we demonstrate how the electric response of MOFs can be tuned by adding rotatable dipolar linkers, generating a material that exhibits paralectric behavior in two dimensions and dielectric behavior in one dimension. The suitability of four different methods to compute the relative permittivity κ by means of molecular dynamics simulations was validated. The dependency of the permittivity on temperature T and dipole strength μ was determined. It was found that the herein investigated systems exhibit a high degree of tunability and substantially larger dielectric constants as expected for MOFs in general. The temperature dependency of κ obeys the Curie-Weiss law. In addition, the influence of dipolar linkers on the electric field induced breathing behavior was investigated. With increasing dipole moment, lower field strength are required to trigger the contraction. These investigations set the stage for an application of such systems as dielectric sensors, order-disorder ferroelectrics or any scenario where movable dipolar fragments respond to external electric fields.

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