scholarly journals An integrative structural model of the full-length gp16 ATPase in bacteriophage phi29 DNA packaging motor

2020 ◽  
Author(s):  
Abdullah F.U.H. Saeed ◽  
Chun Chan ◽  
Hongxin Guan ◽  
Bing Gong ◽  
Peixuan Guo ◽  
...  
Keyword(s):  
Structural Model ◽  
Md Simulations ◽  
Full Length ◽  
Nucleic Acid Binding ◽  
Multiple Sources ◽  
Cellular Functions ◽  
Motor Complex ◽  
Terminal Domain ◽  
Binding Surface ◽  
The Individual

ABSTRACTBiological motors, ubiquitous in living systems, convert chemical energy into different kinds of mechanical motions critical to cellular functions. Most of these biomotors belong to a group of enzymes known as ATPases, which adopt a multi-subunit ring-shaped structure and hydrolyze adenosine triphosphate (ATP) to generate forces. The gene product 16 (gp16), an ATPase in bacteriophage □29, is among the most powerful biomotors known. It can overcome substantial resisting forces from entropic, electrostatic, and DNA bending sources to package double-stranded DNA (dsDNA) into a preformed protein shell (procapsid). Despite numerous studies of the □29 packaging mechanism, a structure of the full-length gp16 is still lacking, let alone that of the packaging motor complex that includes two additional molecular components: a connector gp10 protein and a prohead RNA (pRNA). Here we report the crystal structure of the C-terminal domain of gp16 (gp16-CTD). Structure-based alignment of gp16-CTD with related RNase H-like nuclease domains revealed a nucleic acid binding surface in gp16-CTD, whereas no nuclease activity has been detected for gp16. Subsequent molecular dynamics (MD) simulations showed that this nucleic acid binding surface is likely essential for pRNA binding. Furthermore, our simulations of a full-length gp16 structural model highlighted a dynamic interplay between the N-terminal domain (NTD) and CTD of gp16, which may play a role in driving DNA movement into the procapsid, providing structural support to the previously proposed inchworm model. Lastly, we assembled an atomic structural model of the complete □29 dsDNA packaging motor complex by integrating structural and experimental data from multiple sources. Collectively, our findings provided a refined inchworm-revolution model for dsDNA translocation in bacteriophage □29 and suggested how the individual domains of gp16 work together to power such translocation.ABSTRACT (SHORT)Biological motors, ubiquitous in living systems, convert chemical energy into different kinds of mechanical motions critical to cellular functions. The gene product 16 (gp16) in bacteriophage □29 is among the most powerful biomotors known, which adopts a multi-subunit ring-shaped structure and hydrolyzes ATP to package double-stranded DNA (dsDNA) into a preformed procapsid. Here we report the crystal structure of the C-terminal domain of gp16 (gp16-CTD). Structure-based alignment and molecular dynamics (MD) simulations revealed an essential binding surface of gp16-CTD for prohead RNA (pRNA), a unique component of the motor complex. Furthermore, our simulations highlighted a dynamic interplay between the N-terminal domain (NTD) and CTD of gp16, which may play a role in driving DNA movement into the procapsid. Lastly, we assembled an atomic structural model of the complete □29 dsDNA packaging motor complex by integrating structural and experimental data from multiple sources. Collectively, our findings provided a refined inchworm-revolution model for dsDNA translocation in bacteriophage □29 and suggested how the individual domains of gp16 work together to power such translocation.

scholarly journals Functional Characterization of Tissue Inhibitor of Metalloproteinase-1 (TIMP-1) N- and C-Terminal Domains duringXenopus laevisDevelopment

2014 ◽  
Vol 2014 ◽  
pp. 1-10 ◽  
Author(s):  
M. A. Nieuwesteeg ◽  
J. A. Willson ◽  
M. Cepeda ◽  
M. A. Fox ◽  
S. Damjanovski
Keyword(s):  
Functional Characterization ◽  
Full Length ◽  
Tissue Inhibitor Of Metalloproteinase ◽  
Tissue Inhibitors Of Metalloproteinases ◽  
Mrna Levels ◽  
Ecm Remodeling ◽  
Protein Levels ◽  
Terminal Domain ◽  
The Individual ◽  
Terminal Domains

Extracellular matrix (ECM) remodeling is essential for facilitating developmental processes. ECM remodeling, accomplished by matrix metalloproteinases (MMPs), is regulated by endogenous tissue inhibitors of metalloproteinases (TIMPs). While the TIMP N-terminal domain is involved in inhibition of MMP activity, the C-terminal domain exhibits cell-signaling activity, which is TIMP and cell type dependent. We have previously examined the distinct roles of theXenopus laevisTIMP-2 and -3 C-terminal domains during development and here examined the unique roles of TIMP-1 N- and C-terminal domains in earlyX. laevisembryos. mRNA microinjection was used to overexpress full-length TIMP-1 or its individual N- or C-terminal domains in embryos. Full-length and C-terminal TIMP-1 resulted in increased lethality compared to N-terminal TIMP-1. Overexpression of C-terminal TIMP-1 resulted in significant decreases in mRNA levels of proteolytic genes including TIMP-2, RECK, MMP-2, and MMP-9, corresponding to decreases in MMP-2 and -9 protein levels, as well as decreased MMP-2 and MMP-9 activities. These trends were not observed with the N-terminus. Our research suggests that the individual domains of TIMP-1 are capable of playing distinct roles in regulating the ECM proteolytic network during development and that the unique functions of these domains are moderated in the endogenous full-length TIMP-1 molecule.

scholarly journals Integrative modelling of the full-length human dehydrodolichyl diphosphate synthase using a hybrid computational and experimental approach

2019 ◽  
Author(s):  
Michal Lisnyansky Barel ◽  
Su Youn Lee ◽  
Ah Young Ki ◽  
Noa Kapelushnik ◽  
Anat Loewenstein ◽  
...  
Keyword(s):  
Structural Model ◽  
Clinical Importance ◽  
Protein Glycosylation ◽  
Full Length ◽  
Scattering Data ◽  
Exchange Mass Spectrometry ◽  
Functional Studies ◽  
Terminal Domain ◽  
Helix Loop Helix ◽  
Structural Framework

AbstractDehydrodolichyl diphosphate synthase (DHDDS) and Nogo-B receptor (NgBR) form the heteromeric human cis-prenyltransferase complex, synthesizing the precursor for the glycosyl carrier involved in N-linked protein glycosylation. In line with the important role of N-glycosylation in protein biogenesis, mutations in DHDDS, the catalytic subunit of the complex, were shown to result in human diseases. Importantly, well-characterized DHDDS homologs function as homodimers and not as heteromeric complexes. Moreover, DHDDS encompasses a C-terminal region, which does not converge with any known conserved domains. Therefore, despite the clinical importance of DHDDS, our understating of its structure-function relations remains poor. Here, we provide a structural model for the full-length human DHDDS using a multidisciplinary experimental and computational approach. Our model suggests that the C-terminal domain of DHDDS forms a helix-loop-helix motif, tightly packed against the core catalytic cis-prenyltransferase domain. This model is consistent with small-angle X-ray scattering data, indicating that the full-length DHDDS maintains a similar conformation in solution. Moreover, hydrogen-deuterium exchange mass-spectrometry experiments show time-dependent deuterium uptake in the C-terminal domain, consistent with its overall folded state. Finally, we provide a model for the DHDDS-NgBR heterodimer, offering a structural framework for future structural and functional studies of the human cis-prenyltransferase complex.

scholarly journals Prion-like C-terminal domain of TDP-43 and α-Synuclein interact synergistically to generate neurotoxic hybrid fibrils

2020 ◽  
Author(s):  
Shailendra Dhakal ◽  
Courtney E Wyant ◽  
Hannah E George ◽  
Sarah E Morgan ◽  
Vijayaraghavan Rangachari
Keyword(s):  
Lewy Bodies ◽  
Amyloid Formation ◽  
Liquid Droplets ◽  
Cellular Functions ◽  
Parkinsons Disease ◽  
Terminal Domain ◽  
Amyloid Aggregates ◽  
The Individual ◽  
Lateral Sclerosis ◽  
Selective Interactions

Aberrant aggregation and amyloid formation of tar DNA binding protein (TDP-43) and αsynuclein (αS) underlie frontotemporal dementia (FTD) and Parkinsons disease (PD), respectively. Amyloid inclusions of TDP-43 and αS are also commonly co-observed in amyotrophic lateral sclerosis (ALS), dementia with Lewy bodies (DLB) and Alzheimer disease (AD). Emerging evidence from cellular and animal models show colocalization of the TDP-43 and αS aggregates, raising the possibility of direct interactions and coaggregation between the two proteins. In this report, we set out to answer this question by investigating the interactions between αS and prion-like pathogenic C-terminal domain of TDP-43 (TDP-43 PrLD). PrLD is an aggregation-prone fragment generated both by alternative splicing as well as aberrant proteolytic cleavage of full length TDP-43. Our results indicate that two proteins interact in a synergistic manner to augment each others aggregation towards hybrid fibrils. While monomers, oligomers and sonicated fibrils of αS seed TDP-43 PrLD monomer aggregation, TDP-43 PrLD fibrils failed to seed αS monomers indicating selective interactions. Furthermore, αS modulates liquid droplets formed by TDP-43 PrLD and RNA to promote insoluble amyloid aggregates. Importantly, the cross-seeded hybrid aggregates show greater cytotoxicity as compared to the individual homotypic aggregates suggesting that the interactions between the two proteins have a discernable impact on cellular functions. Together, these results bring forth insights into TDP-43 PrLD - αS interactions that could help explain clinical and pathological presentations in patients with co-morbidities involving the two proteins.

scholarly journals Structure of the full-length human Pannexin1 channel and insights into its role in pyroptosis

2021 ◽  
Vol 7 (1) ◽  
Author(s):  
Sensen Zhang ◽  
Baolei Yuan ◽  
Jordy Homing Lam ◽  
Jun Zhou ◽  
Xuan Zhou ◽  
...  
Keyword(s):  
Md Simulation ◽  
Potential Role ◽  
Md Simulations ◽  
Full Length ◽  
Inflammasome Activation ◽  
Simulation Studies ◽  
Cryo Electron Microscopy ◽  
Gating Mechanism ◽  
Atp Efflux

AbstractPannexin1 (PANX1) is a large-pore ATP efflux channel with a broad distribution, which allows the exchange of molecules and ions smaller than 1 kDa between the cytoplasm and extracellular space. In this study, we show that in human macrophages PANX1 expression is upregulated by diverse stimuli that promote pyroptosis, which is reminiscent of the previously reported lipopolysaccharide-induced upregulation of PANX1 during inflammasome activation. To further elucidate the function of PANX1, we propose the full-length human Pannexin1 (hPANX1) model through cryo-electron microscopy (cryo-EM) and molecular dynamics (MD) simulation studies, establishing hPANX1 as a homo-heptamer and revealing that both the N-termini and C-termini protrude deeply into the channel pore funnel. MD simulations also elucidate key energetic features governing the channel that lay a foundation to understand the channel gating mechanism. Structural analyses, functional characterizations, and computational studies support the current hPANX1-MD model, suggesting the potential role of hPANX1 in pyroptosis during immune responses.

scholarly journals Structural Basis for the C-Terminal Domain of Mycobacterium tuberculosis Ribosome Maturation Factor RimM to Bind Ribosomal Protein S19

Biomolecules ◽  
2021 ◽  
Vol 11 (4) ◽  
pp. 597
Author(s):  
Haoran Zhang ◽  
Qiuxiang Zhou ◽  
Chenyun Guo ◽  
Liubin Feng ◽  
Huilin Wang ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Ribosomal Protein ◽  
Solution Structure ◽  
Molecular Mechanisms ◽  
Structural Model ◽  
Ribosomal Subunit ◽  
Structural Basis ◽  
Hydrophobic Core ◽  
Terminal Domain ◽  
Ribosomal Protein S19

Multidrug-resistant tuberculosis (TB) is a serious threat to public health, calling for the development of new anti-TB drugs. Chaperon protein RimM, involved in the assembly of ribosomal protein S19 into 30S ribosomal subunit during ribosome maturation, is a potential drug target for TB treatment. The C-terminal domain (CTD) of RimM is primarily responsible for binding S19. However, both the CTD structure of RimM from Mycobacterium tuberculosis (MtbRimMCTD) and the molecular mechanisms underlying MtbRimMCTD binding S19 remain elusive. Here, we report the solution structure, dynamics features of MtbRimMCTD, and its interaction with S19. MtbRimMCTD has a rigid hydrophobic core comprised of a relatively conservative six-strand β-barrel, tailed with a short α-helix and interspersed with flexible loops. Using several biophysical techniques including surface plasmon resonance (SPR) affinity assays, nuclear magnetic resonance (NMR) assays, and molecular docking, we established a structural model of the MtbRimMCTD–S19 complex and indicated that the β4-β5 loop and two nonconserved key residues (D105 and H129) significantly contributed to the unique pattern of MtbRimMCTD binding S19, which might be implicated in a form of orthogonality for species-dependent RimM–S19 interaction. Our study provides the structural basis for MtbRimMCTD binding S19 and is beneficial to the further exploration of MtbRimM as a potential target for the development of new anti-TB drugs.

scholarly journals Tethering-induced destabilization and ATP-binding for tandem RRM domains of ALS-causing TDP-43 and hnRNPA1

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Mei Dang ◽  
Yifan Li ◽  
Jianxing Song
Keyword(s):  
Conformational Dynamics ◽  
Md Simulations ◽  
Atp Binding ◽  
Rna Recognition Motif ◽  
Rna Recognition ◽  
Nucleic Acid Binding ◽  
General Role ◽  
Nmr Characterization ◽  
Lateral Sclerosis

AbstractTDP-43 and hnRNPA1 contain tandemly-tethered RNA-recognition-motif (RRM) domains, which not only functionally bind an array of nucleic acids, but also participate in aggregation/fibrillation, a pathological hallmark of various human diseases including amyotrophic lateral sclerosis (ALS), frontotemporal dementia (FTD), alzheimer's disease (AD) and Multisystem proteinopathy (MSP). Here, by DSF, NMR and MD simulations we systematically characterized stability, ATP-binding and conformational dynamics of TDP-43 and hnRNPA1 RRM domains in both tethered and isolated forms. The results reveal three key findings: (1) upon tethering TDP-43 RRM domains become dramatically coupled and destabilized with Tm reduced to only 49 °C. (2) ATP specifically binds TDP-43 and hnRNPA1 RRM domains, in which ATP occupies the similar pockets within the conserved nucleic-acid-binding surfaces, with the affinity slightly higher to the tethered than isolated forms. (3) MD simulations indicate that the tethered RRM domains of TDP-43 and hnRNPA1 have higher conformational dynamics than the isolated forms. Two RRM domains become coupled as shown by NMR characterization and analysis of inter-domain correlation motions. The study explains the long-standing puzzle that the tethered TDP-43 RRM1–RRM2 is particularly prone to aggregation/fibrillation, and underscores the general role of ATP in inhibiting aggregation/fibrillation of RRM-containing proteins. The results also rationalize the observation that the risk of aggregation-causing diseases increases with aging.

scholarly journals Development of an Androgen Receptor Inhibitor Targeting the N-Terminal Domain of Androgen Receptor for Treatment of Castration Resistant Prostate Cancer

Cancers ◽  
2021 ◽  
Vol 13 (14) ◽  
pp. 3488
Author(s):  
Fuqiang Ban ◽  
Eric Leblanc ◽  
Ayse Derya Cavga ◽  
Chia-Chi Flora Huang ◽  
Mark R. Flory ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
Androgen Receptor ◽  
Splice Variants ◽  
Full Length ◽  
Cancer Resistance ◽  
Castration Resistant Prostate Cancer ◽  
Terminal Domain ◽  
Castration Resistant ◽  
Coregulatory Proteins ◽  
Androgen Binding

Prostate cancer patients undergoing androgen deprivation therapy almost invariably develop castration-resistant prostate cancer. Resistance can occur when mutations in the androgen receptor (AR) render anti-androgen drugs ineffective or through the expression of constitutively active splice variants lacking the androgen binding domain entirely (e.g., ARV7). In this study, we are reporting the discovery of a novel AR-NTD covalent inhibitor 1-chloro-3-[(5-([(2S)-3-chloro-2-hydroxypropyl]amino)naphthalen-1-yl)amino]propan-2-ol (VPC-220010) targeting the AR-N-terminal Domain (AR-NTD). VPC-220010 inhibits AR-mediated transcription of full length and truncated variant ARV7, downregulates AR response genes, and selectively reduces the growth of both full-length AR- and truncated AR-dependent prostate cancer cell lines. We show that VPC-220010 disrupts interactions between AR and known coactivators and coregulatory proteins, such as CHD4, FOXA1, ZMIZ1, and several SWI/SNF complex proteins. Taken together, our data suggest that VPC-220010 is a promising small molecule that can be further optimized into effective AR-NTD inhibitor for the treatment of CRPC.

The Entextualization of Ayurveda as Intellectual Property

2012 ◽  
Vol 19 (3) ◽  
pp. 313-343 ◽  
Author(s):  
Matthew Wolfgram
Keyword(s):  
Intellectual Property ◽  
Global Economy ◽  
Medical Knowledge ◽  
Linguistic Anthropology ◽  
Medical Discourse ◽  
Complex Nature ◽  
Multiple Sources ◽  
The Social ◽  
Rights Discourse ◽  
The Individual

AbstractThis article documents the practices of pharmaceutical creativity in Ayurveda, focusing in particular on how practitioners appropriate multiple sources to innovate medical knowledge. Drawing on research in linguistic anthropology on the social circulation of discourse—a process calledentextualization—I describe how the ways in which Ayurveda practitioners innovate medical knowledge confounds the dichotomous logic of intellectual property (IP) rights discourse, which opposes traditional collective knowledge and modern individual innovation. While it is clear that these categories do not comprehend the complex nature of creativity in Ayurveda, I also use the concept of entextualization to describe how recent historical shifts in the circulation of discourse have caused a partial entailment of this opposition between the individual and the collectivity. Ultimately, I argue that the method exemplified in this article of tracking the social circulation of medical discourse highlights both the empirical complexity of so-called traditional creativity, and the politics of imposing the categories of IP rights discourse upon that creativity, situated as it often is, at the margins of the global economy.

scholarly journals ADAMTS13 and 15 are not regulated by the full length and N-terminal domain forms of TIMP-1, -2, -3 and -4

2015 ◽  
Vol 4 (1) ◽  
pp. 73-78 ◽  
Author(s):  
CENQI GUO ◽  
ANASTASIA TSIGKOU ◽  
MENG HUEE LEE
Keyword(s):  
Full Length ◽  
Terminal Domain

scholarly journals Molecular insights on CALX-CBD12 inter-domain dynamics from MD simulations, RDCs and SAXS

2020 ◽  
Author(s):  
Maximilia F. de Souza Degenhardt ◽  
Phelipe A. M. Vitale ◽  
Layara A. Abiko ◽  
Martin Zacharias ◽  
Michael Sattler ◽  
...  
Keyword(s):  
Transmembrane Domain ◽  
Md Simulations ◽  
Bound State ◽  
Conformational Ensemble ◽  
Regulation Mechanism ◽  
Multiple Sources ◽  
Intracellular Loop ◽  
Saxs Data ◽  
Structure Information ◽  
Apo State

ABSTRACTNa+/Ca2+ exchangers (NCX) are secondary active transporters that couple the translocation of Na+ with the transport of Ca2+ in the opposite direction. The exchanger is an essential Ca2+ extrusion mechanism in excitable cells. It consists of a transmembrane domain and a large intracellular loop that contains two Ca2+-binding domains, CBD1 and CBD2. The two CBDs are adjacent to each other and form a two-domain Ca2+-sensor called CBD12. Binding of intracellular Ca2+ to CBD12 activates the NCX but inhibits the Na+/Ca2+ exchanger of Drosophila, CALX. NMR spectroscopy and SAXS studies showed that CALX and NCX CBD12 constructs display significant inter-domain flexibility in the Apo state, but assume rigid inter-domain arrangements in the Ca2+-bound state. However, detailed structure information on CBD12 in the Apo state is missing. Structural characterization of proteins formed by two or more domains connected by flexible linkers is notoriously challenging and requires the combination of orthogonal information from multiple sources. As an attempt to characterize the conformational ensemble of CALX-CBD12 in the Apo state, we applied molecular dynamics (MD) simulations, NMR (1H-15N RDCs) and Small-Angle X-Ray Scattering (SAXS) data in a combined modelling strategy that generated atomistic information on the most representative conformations. This joint approach demonstrated that CALX-CBD12 preferentially samples closed conformations, while the wide-open inter-domain arrangement characteristic of the Ca2+-bound state is less frequently sampled. These results are consistent with the view that Ca2+ binding shifts the CBD12 conformational ensemble towards extended conformers, which could be a key step in the Na+/Ca2+ exchangers’ allosteric regulation mechanism. The present strategy, combining MD with NMR and SAXS, provides a powerful approach to select representative structures from ensembles of conformations, which could be applied to other flexible multi-domain systems.SIGNIFICANCEThe conformational ensemble of CALX-CBD12, the main Ca2+-sensor of Drosophila’s Na+/Ca2+ exchanger, was characterized by a combination of MD simulations with SAXS and NMR data using the EOM approach. This analysis showed that this two-domain construct experiences opening-closing motions, providing molecular information about CALX-CBD12 in the Apo state. Ca2+-binding shifts the conformational ensemble towards extended conformers. These findings are consistent with a model according to which Ca2+ modulation of CBD12 plasticity is a key step in the Ca2+-regulation mechanism of the full-length exchanger.

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