scholarly journals A single amino acid polymorphism in a conserved effector of the multihost blast fungus pathogen expands host-target binding spectrum

2021 ◽  
Vol 17 (11) ◽  
pp. e1009957
Author(s):  
Adam R. Bentham ◽  
Yohann Petit-Houdenot ◽  
Joe Win ◽  
Izumi Chuma ◽  
Ryohei Terauchi ◽  
...  
Keyword(s):  
Structural Biology ◽  
Adaptive Evolution ◽  
Host Specialization ◽  
Single Amino Acid ◽  
Structural Basis ◽  
Common Mechanism ◽  
Ancestral Reconstruction ◽  
Blast Fungus ◽  
Pathogen Effector ◽  
Binding Interfaces

Accelerated gene evolution is a hallmark of pathogen adaptation and specialization following host-jumps. However, the molecular processes associated with adaptive evolution between host-specific lineages of a multihost plant pathogen remain poorly understood. In the blast fungus Magnaporthe oryzae (Syn. Pyricularia oryzae), host specialization on different grass hosts is generally associated with dynamic patterns of gain and loss of virulence effector genes that tend to define the distinct genetic lineages of this pathogen. Here, we unravelled the biochemical and structural basis of adaptive evolution of APikL2, an exceptionally conserved paralog of the well-studied rice-lineage specific effector AVR-Pik. Whereas AVR-Pik and other members of the six-gene AVR-Pik family show specific patterns of presence/absence polymorphisms between grass-specific lineages of M. oryzae, APikL2 stands out by being ubiquitously present in all blast fungus lineages from 13 different host species. Using biochemical, biophysical and structural biology methods, we show that a single aspartate to asparagine polymorphism expands the binding spectrum of APikL2 to host proteins of the heavy-metal associated (HMA) domain family. This mutation maps to one of the APikL2-HMA binding interfaces and contributes to an altered hydrogen-bonding network. By combining phylogenetic ancestral reconstruction with an analysis of the structural consequences of allelic diversification, we revealed a common mechanism of effector specialization in the AVR-Pik/APikL2 family that involves two major HMA-binding interfaces. Together, our findings provide a detailed molecular evolution and structural biology framework for diversification and adaptation of a fungal pathogen effector family following host-jumps.

scholarly journals A single amino acid polymorphism in a conserved effector of the multihost blast fungus pathogen expands host-target binding spectrum

2021 ◽  
Author(s):  
Adam R. Bentham ◽  
Yohann Petit-Houdenot ◽  
Joe Win ◽  
Izumi Chuma ◽  
Ryohei Terauchi ◽  
...  
Keyword(s):  
Structural Biology ◽  
Adaptive Evolution ◽  
Host Specialization ◽  
Single Amino Acid ◽  
Structural Basis ◽  
Common Mechanism ◽  
Ancestral Reconstruction ◽  
Blast Fungus ◽  
Pathogen Effector ◽  
Binding Interfaces

AbstractAccelerated gene evolution is a hallmark of pathogen adaptation and specialization following host-jumps. However, the molecular processes associated with adaptive evolution between host-specific lineages of a multihost plant pathogen remain poorly understood. In the blast fungus Magnaporthe oryzae (Syn. Pyricularia oryzae), host specialization on different grass hosts is generally associated with dynamic patterns of gain and loss of virulence effector genes that tend to define the distinct genetic lineages of this pathogen. Here, we unravelled the biochemical and structural basis of adaptive evolution of APikL2, an exceptionally conserved paralog of the well-studied rice-lineage specific effector AVR-Pik. Whereas AVR-Pik and other members of the six-gene AVR-Pik family show specific patterns of presence/absence polymorphisms between grassspecific lineages of M. oryzae, APikL2 stands out by being ubiquitously present in all blast fungus lineages from 13 different host species. Using biochemical, biophysical and structural biology methods, we show that a single aspartate to asparagine polymorphism expands the binding spectrum of APikL2 to host proteins of the heavy-metal associated (HMA) domain family. This mutation maps to one of the APikL2-HMA binding interfaces and contributes to an altered hydrogen-bonding network. By combining phylogenetic ancestral reconstruction with an analysis of the structural consequences of allelic diversification, we revealed a common mechanism of effector specialization in the AVR-Pik/APikL2 family that involves two major HMA-binding interfaces. Together, our findings provide a detailed molecular evolution and structural biology framework for diversification and adaptation of a fungal pathogen effector family following host-jumps.

scholarly journals Structural Basis for Linezolid Binding Site Rearrangement in the Staphylococcus aureus Ribosome

mBio ◽  
2017 ◽  
Vol 8 (3) ◽  
Author(s):  
Matthew J. Belousoff ◽  
Zohar Eyal ◽  
Mazdak Radjainia ◽  
Tofayel Ahmed ◽  
Rebecca S. Bamert ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Structural Information ◽  
Structural Basis ◽  
Common Mechanism ◽  
Resistance Mutations ◽  
Antimicrobial Drugs ◽  
Linezolid Resistance ◽  
Drug Modification ◽  
Antibiotic Resistance Mutations ◽  
Shed Light

ABSTRACT An unorthodox, surprising mechanism of resistance to the antibiotic linezolid was revealed by cryo-electron microscopy (cryo-EM) in the 70S ribosomes from a clinical isolate of Staphylococcus aureus. This high-resolution structural information demonstrated that a single amino acid deletion in ribosomal protein uL3 confers linezolid resistance despite being located 24 Å away from the linezolid binding pocket in the peptidyl-transferase center. The mutation induces a cascade of allosteric structural rearrangements of the rRNA that ultimately results in the alteration of the antibiotic binding site. IMPORTANCE The growing burden on human health caused by various antibiotic resistance mutations now includes prevalent Staphylococcus aureus resistance to last-line antimicrobial drugs such as linezolid and daptomycin. Structure-informed drug modification represents a frontier with respect to designing advanced clinical therapies, but success in this strategy requires rapid, facile means to shed light on the structural basis for drug resistance (D. Brown, Nat Rev Drug Discov 14:821–832, 2015, https://doi.org/10.1038/nrd4675 ). Here, detailed structural information demonstrates that a common mechanism is at play in linezolid resistance and provides a step toward the redesign of oxazolidinone antibiotics, a strategy that could thwart known mechanisms of linezolid resistance. IMPORTANCE The growing burden on human health caused by various antibiotic resistance mutations now includes prevalent Staphylococcus aureus resistance to last-line antimicrobial drugs such as linezolid and daptomycin. Structure-informed drug modification represents a frontier with respect to designing advanced clinical therapies, but success in this strategy requires rapid, facile means to shed light on the structural basis for drug resistance (D. Brown, Nat Rev Drug Discov 14:821–832, 2015, https://doi.org/10.1038/nrd4675 ). Here, detailed structural information demonstrates that a common mechanism is at play in linezolid resistance and provides a step toward the redesign of oxazolidinone antibiotics, a strategy that could thwart known mechanisms of linezolid resistance.

scholarly journals The Cruciality of Single Amino Acid Replacement for the Spectral Tuning of Biliverdin-Binding Cyanobacteriochromes

2020 ◽  
Vol 21 (17) ◽  
pp. 6278
Author(s):  
Keiji Fushimi ◽  
Hiroki Hoshino ◽  
Naeko Shinozaki-Narikawa ◽  
Yuto Kuwasaki ◽  
Keita Miyake ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Molecular Mechanisms ◽  
Near Infrared ◽  
Structural Characteristics ◽  
Blue Shift ◽  
Amino Acid Replacement ◽  
Red Shift ◽  
Single Amino Acid ◽  
Structural Basis ◽  
Small Unit

Cyanobacteriochromes (CBCRs), which are known as linear tetrapyrrole-binding photoreceptors, to date can only be detected from cyanobacteria. They can perceive light only in a small unit, which is categorized into various lineages in correlation with their spectral and structural characteristics. Recently, we have succeeded in identifying specific molecules, which can incorporate mammalian intrinsic biliverdin (BV), from the expanded red/green (XRG) CBCR lineage and in converting BV-rejective molecules into BV-acceptable ones with the elucidation of the structural basis. Among the BV-acceptable molecules, AM1_1870g3_BV4 shows a spectral red-shift in comparison with other molecules, while NpF2164g5_BV4 does not show photoconversion but stably shows a near-infrared (NIR) fluorescence. In this study, we found that AM1_1870g3_BV4 had a specific Tyr residue near the d-ring of the chromophore, while others had a highly conserved Leu residue. The replacement of this Tyr residue with Leu in AM1_1870g3_BV4 resulted in a blue-shift of absorption peak. In contrast, reverse replacement in NpF2164g5_BV4 resulted in a red-shift of absorption and fluorescence peaks, which applies to fluorescence bio-imaging in mammalian cells. Notably, the same Tyr/Leu-dependent color-tuning is also observed for the CBCRs belonging to the other lineage, which indicates common molecular mechanisms.

scholarly journals Structural Characterization of the Helicase nsp10 Encoded by Porcine Reproductive and Respiratory Syndrome Virus

2020 ◽  
Vol 94 (15) ◽  
Author(s):  
Yuejun Shi ◽  
Xiaohan Tong ◽  
Gang Ye ◽  
Ruixue Xiu ◽  
Lisha Li ◽  
...  
Keyword(s):  
Domain Structure ◽  
Crucial Role ◽  
Catalytic Domain ◽  
Full Length ◽  
Structural Basis ◽  
Economic Losses ◽  
Respiratory Syndrome Virus ◽  
Common Mechanism ◽  
Helicase Activity ◽  
Terminal Domain

ABSTRACT Currently, an effective therapeutic treatment for porcine reproductive and respiratory syndrome virus (PRRSV) remains elusive. PRRSV helicase nsp10 is an important component of the replication transcription complex that plays a crucial role in viral replication, making nsp10 an important target for drug development. Here, we report the first crystal structure of full-length nsp10 from the arterivirus PRRSV, which has multiple domains: an N-terminal zinc-binding domain (ZBD), a 1B domain, and helicase core domains 1A and 2A. Importantly, our structural analyses indicate that the conformation of the 1B domain from arterivirus nsp10 undergoes a dynamic transition. The polynucleotide substrate channel formed by domains 1A and 1B adopts an open state, which may create enough space to accommodate and bind double-stranded RNA (dsRNA) during unwinding. Moreover, we report a unique C-terminal domain structure that participates in stabilizing the overall helicase structure. Our biochemical experiments also showed that deletion of the 1B domain and C-terminal domain significantly reduced the helicase activity of nsp10, indicating that the four domains must cooperate to contribute to helicase function. In addition, our results indicate that nidoviruses contain a conserved helicase core domain and key amino acid sites affecting helicase function, which share a common mechanism of helicase translocation and unwinding activity. These findings will help to further our understanding of the mechanism of helicase function and provide new targets for the development of antiviral drugs. IMPORTANCE Porcine reproductive and respiratory syndrome virus (PRRSV) is a major respiratory disease agent in pigs that causes enormous economic losses to the global swine industry. PRRSV helicase nsp10 is a multifunctional protein with translocation and unwinding activities and plays a vital role in viral RNA synthesis. Here, we report the first structure of full-length nsp10 from the arterivirus PRRSV at 3.0-Å resolution. Our results show that the 1B domain of PRRSV nsp10 adopts a novel open state and has a unique C-terminal domain structure, which plays a crucial role in nsp10 helicase activity. Furthermore, mutagenesis and structural analysis revealed conservation of the helicase catalytic domain across the order Nidovirales (families Arteriviridae and Coronaviridae). Importantly, our results will provide a structural basis for further understanding the function of helicases in the order Nidovirales.

scholarly journals Structural basis for GPCR-independent activation of heterotrimeric Gi proteins

2019 ◽  
Vol 116 (33) ◽  
pp. 16394-16403 ◽  
Author(s):  
Nicholas A. Kalogriopoulos ◽  
Steven D. Rees ◽  
Tony Ngo ◽  
Noah J. Kopcho ◽  
Andrey V. Ilatovskiy ◽  
...  
Keyword(s):  
G Protein ◽  
G Proteins ◽  
Structural Changes ◽  
Control Cell ◽  
Structural Basis ◽  
Common Mechanism ◽  
Hydrophobic Core ◽  
Nucleotide Exchange ◽  
Protein Activation ◽  
G Protein Activation

Heterotrimeric G proteins are key molecular switches that control cell behavior. The canonical activation of G proteins by agonist-occupied G protein-coupled receptors (GPCRs) has recently been elucidated from the structural perspective. In contrast, the structural basis for GPCR-independent G protein activation by a novel family of guanine-nucleotide exchange modulators (GEMs) remains unknown. Here, we present a 2.0-Å crystal structure of Gαi in complex with the GEM motif of GIV/Girdin. Nucleotide exchange assays, molecular dynamics simulations, and hydrogen–deuterium exchange experiments demonstrate that GEM binding to the conformational switch II causes structural changes that allosterically propagate to the hydrophobic core of the Gαi GTPase domain. Rearrangement of the hydrophobic core appears to be a common mechanism by which GPCRs and GEMs activate G proteins, although with different efficiency. Atomic-level insights presented here will aid structure-based efforts to selectively target the noncanonical G protein activation.

Molecular mousetraps and the serpinopathies1

2005 ◽  
Vol 33 (2) ◽  
pp. 321-330 ◽  
Author(s):  
D.A. Lomas ◽  
D. Belorgey ◽  
M. Mallya ◽  
E. Miranda ◽  
K.J. Kinghorn ◽  
...  
Keyword(s):  
Conformational Transition ◽  
Point Mutations ◽  
Serine Proteinase ◽  
Structural Basis ◽  
Common Mechanism ◽  
Secretory Cells ◽  
C1 Inhibitor ◽  
Serine Proteinase Inhibitor ◽  
Clinical Syndromes ◽  
The Common

Members of the serine proteinase inhibitor or serpin superfamily inhibit their target proteinases by a remarkable conformational transition that involves the enzyme being translocated more than 70 Å (1 Å=10−10 m) from the upper to the lower pole of the inhibitor. This elegant mechanism is subverted by point mutations to form ordered polymers that are retained within the endoplasmic reticulum of secretory cells. The accumulation of polymers underlies the retention of mutants of α1-antitrypsin and neuroserpin within hepatocytes and neurons to cause cirrhosis and dementia respectively. The formation of polymers results in the failure to secrete mutants of other members of the serpin superfamily: antithrombin, C1 inhibitor and α1-antichymotrypsin, to cause a plasma deficiency that results in the clinical syndromes of thrombosis, angio-oedema and emphysema respectively. Understanding the common mechanism underlying the retention and deficiency of mutants of the serpins has allowed us to group these conditions as the serpinopathies. We review in this paper the molecular and structural basis of the serpinopathies and show how this has allowed the development of specific agents to block the polymerization that underlies disease.

scholarly journals Determination of the structural basis for selective binding of Epstein-Barr virus to human complement receptor type 2.

1991 ◽  
Vol 174 (6) ◽  
pp. 1299-1311 ◽  
Author(s):  
D R Martin ◽  
A Yuryev ◽  
K R Kalli ◽  
D T Fearon ◽  
J M Ahearn
Keyword(s):  
Epstein Barr Virus ◽  
Specific Binding ◽  
Glycosylation Site ◽  
Single Amino Acid ◽  
Structural Basis ◽  
Viral Receptor ◽  
Barr Virus ◽  
Preferential Binding ◽  
Epstein Barr ◽  
Species Specific

Epstein-Barr virus (EBV) is an oncogenic herpesvirus that selectively infects and immortalizes human B lymphocytes. One determinant of this narrow tropism is human CR2, the only viral receptor within the superfamily of proteins that contain short consensus repeats (SCRs). Human CR2 serves as a receptor for both C3dg and the gp350/220 glycoprotein of EBV, and binds the monoclonal antibody (mAb) OKB7, which blocks binding of both ligands to the receptor. In contrast, although murine CR2 is capable of binding human C3dg and this interaction can be blocked with the mAb 7G6, it does not bind OKB7 or EBV. We have determined the structural basis for absolute specificity of EBV for human CR2 through characterization of a panel of 24 human-murine chimeric receptors, all of which bind human C3dg. The results indicate that preferential binding of EBV to human CR2 is not due to unique amino acids that are capable of binding the virus, but reflects a distinct receptor conformation that can be achieved in murine CR2 with single amino acid substitutions in two discontinuous regions of the primary structure: replacement of proline at position 15 with the corresponding serine from human CR2, and elimination of a potential N-linked glycosylation site between SCR-1 and SCR-2. Furthermore, species-specific binding of EBV, OKB7, and 7G6 can all be manipulated through substitutions among residues 8-15, suggesting that this octapeptide is part of a structural determinant that is critical for binding of both viral and natural ligands to CR2.

scholarly journals Crystal Structure and Structural Mechanism of a Novel Anti-Human Immunodeficiency Virus and d-Amino Acid-Containing Chemokine

2007 ◽  
Vol 81 (20) ◽  
pp. 11489-11498 ◽  
Author(s):  
Dongxiang Liu ◽  
Navid Madani ◽  
Ying Li ◽  
Rong Cao ◽  
Won-Tak Choi ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Human Immunodeficiency Virus ◽  
Amino Acid ◽  
Structural Biology ◽  
Chemokine Receptors ◽  
Structural Basis ◽  
Receptor Selectivity ◽  
Wide Range ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT Chemokines and their receptors play important roles in normal physiological functions and the pathogeneses of a wide range of human diseases, including the entry of human immunodeficiency virus type 1 (HIV-1). However, the use of natural chemokines to probe receptor biology or to develop therapeutic drugs is limited by their lack of selectivity and the poor understanding of mechanisms in ligand-receptor recognition. We addressed these issues by combining chemical and structural biology in research into molecular recognition and inhibitor design. Specifically, the concepts of chemical biology were used to develop synthetically and modularly modified (SMM) chemokines that are unnatural and yet have properties improved over those of natural chemokines in terms of receptor selectivity, affinity, and the ability to explore receptor functions. This was followed by using structural biology to determine the structural basis for synthetically perturbed ligand-receptor selectivity. As a proof-of-principle for this combined chemical and structural-biology approach, we report a novel d-amino acid-containing SMM-chemokine designed based on the natural chemokine called viral macrophage inflammatory protein II (vMIP-II). The incorporation of unnatural d-amino acids enhanced the affinity of this molecule for CXCR4 but significantly diminished that for CCR5 or CCR2, thus yielding much more selective recognition of CXCR4 than wild-type vMIP-II. This d-amino acid-containing chemokine also showed more potent and specific inhibitory activity against HIV-1 entry via CXCR4 than natural chemokines. Furthermore, the high-resolution crystal structure of this d-amino acid-containing chemokine and a molecular-modeling study of its complex with CXCR4 provided the structure-based mechanism for the selective interaction between the ligand and chemokine receptors and the potent anti-HIV activity of d-amino acid-containing chemokines.

scholarly journals Elucidation of AMPA receptor–stargazin complexes by cryo–electron microscopy

Science ◽  
2016 ◽  
Vol 353 (6294) ◽  
pp. 83-86 ◽  
Author(s):  
Edward C. Twomey ◽  
Maria V. Yelshanskaya ◽  
Robert A. Grassucci ◽  
Joachim Frank ◽  
Alexander I. Sobolevsky
Keyword(s):  
Electron Microscopy ◽  
Cognitive Processes ◽  
Electrostatic Interactions ◽  
Structural Basis ◽  
Cryo Electron Microscopy ◽  
Activated State ◽  
Ampar Trafficking ◽  
Excitatory Neurotransmission ◽  
Binding Interfaces ◽  
The Brain

AMPA-subtype ionotropic glutamate receptors (AMPARs) mediate fast excitatory neurotransmission and contribute to high cognitive processes such as learning and memory. In the brain, AMPAR trafficking, gating, and pharmacology is tightly controlled by transmembrane AMPAR regulatory proteins (TARPs). Here, we used cryo–electron microscopy to elucidate the structural basis of AMPAR regulation by one of these auxiliary proteins, TARP γ2, or stargazin (STZ). Our structures illuminate the variable interaction stoichiometry of the AMPAR-TARP complex, with one or two TARP molecules binding one tetrameric AMPAR. Analysis of the AMPAR-STZ binding interfaces suggests that electrostatic interactions between the extracellular domains of AMPAR and STZ play an important role in modulating AMPAR function through contact surfaces that are conserved across AMPARs and TARPs. We propose a model explaining how TARPs stabilize the activated state of AMPARs and how the interactions between AMPARs and their auxiliary proteins control fast excitatory synaptic transmission.

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