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1.
Curr Opin Struct Biol ; 68: 166-174, 2021 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-33540375

RESUMO

Peptidoglycan (PG) is the major structural polymer of the bacterial cell wall. The PG layer of gram-positive bacterial pathogens such as Staphylococcus aureus (S. aureus) is permeated with anionic glycopolymers known as wall teichoic acids (WTAs) and lipoteichoic acids (LTAs). In S. aureus, the WTA backbone typically consists of repeating ribitol-5-phosphate units, which are modified by enzymes that introduce glycosylation as well as amino acids at different locations. These modifications are key determinants of phage adhesion, bacterial biofilm formation and virulence of S. aureus. In this review, we examine differences in WTA structures in gram-positive bacteria, focusing in particular on three enzymes, TarM, TarS, and TarP that glycosylate the WTA of S. aureus at different locations. Infections with S. aureus pose an increasing threat to human health, particularly through the emergence of multidrug-resistant strains. Recently obtained structural information on TarM, TarS and TarP has helped to better understand the strategies used by S. aureus to establish resistance and to evade host defense mechanisms. Moreover, structures of complexes with poly-RboP and its analogs can serve as a platform for the development of new inhibitors that could form a basis for the development of antibiotic agents.

2.
Proc Natl Acad Sci U S A ; 118(3)2021 01 19.
Artigo em Inglês | MEDLINE | ID: mdl-33384338

RESUMO

Human adenovirus species D (HAdV-D) types are currently being explored as vaccine vectors for coronavirus disease 2019 (COVID-19) and other severe infectious diseases. The efficacy of such vector-based vaccines depends on functional interactions with receptors on host cells. Adenoviruses of different species are assumed to enter host cells mainly by interactions between the knob domain of the protruding fiber capsid protein and cellular receptors. Using a cell-based receptor-screening assay, we identified CD46 as a receptor for HAdV-D56. The function of CD46 was validated in infection experiments using cells lacking and overexpressing CD46, and by competition infection experiments using soluble CD46. Remarkably, unlike HAdV-B types that engage CD46 through interactions with the knob domain of the fiber protein, HAdV-D types infect host cells through a direct interaction between CD46 and the hexon protein. Soluble hexon proteins (but not fiber knob) inhibited HAdV-D56 infection, and surface plasmon analyses demonstrated that CD46 binds to HAdV-D hexon (but not fiber knob) proteins. Cryoelectron microscopy analysis of the HAdV-D56 virion-CD46 complex confirmed the interaction and showed that CD46 binds to the central cavity of hexon trimers. Finally, soluble CD46 inhibited infection by 16 out of 17 investigated HAdV-D types, suggesting that CD46 is an important receptor for a large group of adenoviruses. In conclusion, this study identifies a noncanonical entry mechanism used by human adenoviruses, which adds to the knowledge of adenovirus biology and can also be useful for development of adenovirus-based vaccine vectors.


Assuntos
Adenovírus Humanos , Proteínas do Capsídeo , Regulação Viral da Expressão Gênica , Internalização do Vírus , Adenovírus Humanos/genética , Adenovírus Humanos/metabolismo , /metabolismo , Proteínas do Capsídeo/biossíntese , Proteínas do Capsídeo/genética , Linhagem Celular , Humanos
3.
J Mol Biol ; 2020 Nov 26.
Artigo em Inglês | MEDLINE | ID: mdl-33249189

RESUMO

Prenylation is a process widely prevalent in primary and secondary metabolism, contributing to functionality and chemical diversity in natural systems. Due to their high regio- and chemoselectivities, prenyltransferases are also valuable tools for creation of new compounds by chemoenzymatic synthesis and synthetic biology. Over the last ten years, biochemical and structural investigations shed light on the mechanism and key residues that control the catalytic process, but to date crucial information on how certain prenyltransferases control regioselectivity and chemoselectivity was still lacking. Here, we advance a general understanding of the enzyme family by contributing the first structure of a tryptophan C5-prenyltransferase 5-DMATS. Additinallyi, the structure of a bacterial tryptophan C6-prenyltransferase 6-DMATS was solved. Analysis and comparison of both substrate-bound complexes led to the identification of key residues for catalysis. Next, site-directed mutagenesis was successfully implemented to not only modify the prenyl donor specificity but also to redirect the prenylation, thereby switching the regioselectivity of 6-DMATS to that of 5-DMATS. The general strategy of structure-guided protein engineering should be applicable to other related prenyltransferases, thus enabling the production of novel prenylated compounds.

4.
J Virol ; 94(23)2020 Nov 09.
Artigo em Inglês | MEDLINE | ID: mdl-32938765

RESUMO

Reovirus attachment protein σ1 is a trimeric molecule containing tail, body, and head domains. During infection, σ1 engages sialylated glycans and junctional adhesion molecule-A (JAM-A), triggering uptake into the endocytic compartment, where virions are proteolytically converted to infectious subvirion particles (ISVPs). Further disassembly allows σ1 release and escape of transcriptionally active reovirus cores into the cytosol. Electron microscopy has revealed a distinct conformational change in σ1 from a compact form on virions to an extended form on ISVPs. To determine the importance of σ1 conformational mobility, we used reverse genetics to introduce cysteine mutations that can cross-link σ1 by establishing disulfide bonds between structurally adjacent sites in the tail, body, and head domains. We detected phenotypic differences among the engineered viruses. A mutant with a cysteine pair in the head domain replicates with enhanced kinetics, forms large plaques, and displays increased avidity for JAM-A relative to the parental virus, mimicking properties of ISVPs. However, unlike ISVPs, particles containing cysteine mutations that cross-link the head domain uncoat and transcribe viral positive-sense RNA with kinetics similar to the parental virus and are sensitive to ammonium chloride, which blocks virion-to-ISVP conversion. Together, these data suggest that σ1 conformational flexibility modulates the efficiency of reovirus host cell attachment.IMPORTANCE Nonenveloped virus entry is an incompletely understood process. For reovirus, the functional significance of conformational rearrangements in the attachment protein, σ1, that occur during entry and particle uncoating are unknown. We engineered and characterized reoviruses containing cysteine mutations that cross-link σ1 monomers in nonreducing conditions. We found that the introduction of a cysteine pair in the receptor-binding domain of σ1 yielded a virus that replicates with faster kinetics than the parental virus and forms larger plaques. Using functional assays, we found that cross-linking the σ1 receptor-binding domain modulates reovirus attachment but not uncoating or transcription. These data suggest that σ1 conformational rearrangements mediate the efficiency of reovirus host cell binding.

5.
ACS Chem Biol ; 15(10): 2683-2691, 2020 10 16.
Artigo em Inglês | MEDLINE | ID: mdl-32845119

RESUMO

Coxsackievirus A24 variant (CVA24v) and human adenovirus 37 (HAdV-37) are leading causative agents of the severe and highly contagious ocular infections acute hemorrhagic conjunctivitis and epidemic keratoconjunctivitis, respectively. Currently, neither vaccines nor antiviral agents are available for treating these diseases, which affect millions of individuals worldwide. CVA24v and HAdV-37 utilize sialic acid as attachment receptors facilitating entry into host cells. Previously, we and others have shown that derivatives based on sialic acid are effective in preventing HAdV-37 binding and infection of cells. Here, we designed and synthesized novel pentavalent sialic acid conjugates and studied their inhibitory effect against CVA24v and HAdV-37 binding and infection of human corneal epithelial cells. The pentavalent conjugates are the first reported inhibitors of CVA24v infection and proved efficient in blocking HAdV-37 binding. Taken together, the pentavalent conjugates presented here form a basis for the development of general inhibitors of these highly contagious ocular pathogens.

6.
mBio ; 11(4)2020 07 28.
Artigo em Inglês | MEDLINE | ID: mdl-32723915

RESUMO

Asymptomatic infections with polyomaviruses in humans are common, but these small viruses can cause severe diseases in immunocompromised hosts. New Jersey polyomavirus (NJPyV) was identified via a muscle biopsy in an organ transplant recipient with systemic vasculitis, myositis, and retinal blindness, and human polyomavirus 12 (HPyV12) was detected in human liver tissue. The evolutionary origins and potential diseases are not well understood for either virus. In order to define their receptor engagement strategies, we first used nuclear magnetic resonance (NMR) spectroscopy to establish that the major capsid proteins (VP1) of both viruses bind to sialic acid in solution. We then solved crystal structures of NJPyV and HPyV12 VP1 alone and in complex with sialylated glycans. NJPyV employs a novel binding site for a α2,3-linked sialic acid, whereas HPyV12 engages terminal α2,3- or α2,6-linked sialic acids in an exposed site similar to that found in Trichodysplasia spinulosa-associated polyomavirus (TSPyV). Gangliosides or glycoproteins, featuring in mammals usually terminal sialic acids, are therefore receptor candidates for both viruses. Structural analyses show that the sialic acid-binding site of NJPyV is conserved in chimpanzee polyomavirus (ChPyV) and that the sialic acid-binding site of HPyV12 is widely used across the entire polyomavirus family, including mammalian and avian polyomaviruses. A comparison with other polyomavirus-receptor complex structures shows that their capsids have evolved to generate several physically distinct virus-specific receptor-binding sites that can all specifically engage sialylated glycans through a limited number of contacts. Small changes in each site may have enabled host-switching events during the evolution of polyomaviruses.IMPORTANCE Virus attachment to cell surface receptors is critical for productive infection. In this study, we have used a structure-based approach to investigate the cell surface recognition event for New Jersey polyomavirus (NJPyV) and human polyomavirus 12 (HPyV12). These viruses belong to the polyomavirus family, whose members target different tissues and hosts, including mammals, birds, fish, and invertebrates. Polyomaviruses are nonenveloped viruses, and the receptor-binding site is located in their capsid protein VP1. The NJPyV capsid features a novel sialic acid-binding site that is shifted in comparison to other structurally characterized polyomaviruses but shared with a closely related simian virus. In contrast, HPyV12 VP1 engages terminal sialic acids in a manner similar to the human Trichodysplasia spinulosa-associated polyomavirus. Our structure-based phylogenetic analysis highlights that even distantly related avian polyomaviruses possess the same exposed sialic acid-binding site. These findings complement phylogenetic models of host-virus codivergence and may also reflect past host-switching events.

7.
J Virol ; 94(20)2020 09 29.
Artigo em Inglês | MEDLINE | ID: mdl-32699083

RESUMO

Merkel cell polyomavirus (MCPyV) is a human double-stranded DNA tumor virus. MCPyV cell entry is unique among members of the polyomavirus family as it requires the engagement of two types of glycans, sialylated oligosaccharides and sulfated glycosaminoglycans (GAGs). Here, we present crystallographic and cryo-electron microscopic structures of the icosahedral MCPyV capsid and analysis of its glycan interactions via nuclear magnetic resonance (NMR) spectroscopy. While sialic acid binding is specific for α2-3-linked sialic acid and mediated by the exposed apical loops of the major capsid protein VP1, a broad range of GAG oligosaccharides bind to recessed regions between VP1 capsomers. Individual VP1 capsomers are tethered to one another by an extensive disulfide network that differs in architecture from previously described interactions for other PyVs. An unusual C-terminal extension in MCPyV VP1 projects from the recessed capsid regions. Mutagenesis experiments show that this extension is dispensable for receptor interactions.IMPORTANCE The MCPyV genome was found to be clonally integrated in 80% of cases of Merkel cell carcinoma (MCC), a rare but aggressive form of human skin cancer, strongly suggesting that this virus is tumorigenic. In the metastasizing state, the course of the disease is often fatal, especially in immunocompromised individuals, as reflected by the high mortality rate of 33 to 46% and the low 5-year survival rate (<45%). The high seroprevalence of about 60% makes MCPyV a serious health care burden and illustrates the need for targeted treatments. In this study, we present the first high-resolution structural data for this human tumor virus and demonstrate that the full capsid is required for the essential interaction with its GAG receptor(s). Together, these data can be used as a basis for future strategies in drug development.

8.
Med Microbiol Immunol ; 209(3): 325-333, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31784892

RESUMO

Human Adenoviruses (HAdVs) are a family of clinically and therapeutically relevant viruses. A precise understanding of their host cell attachment and entry mechanisms can be applied in inhibitor design and the construction of targeted gene delivery vectors. In this article, structural data on adenovirus attachment and entry are reviewed. HAdVs engage two types of receptors: first, an attachment receptor that is bound by the fibre knob protein protruding from the icosahedral capsid, and next, an integrin entry receptor bound by the pentameric penton base at the capsid vertices. Adenoviruses use remarkably diverse attachment receptors, five of which have been studied structurally in the context of HAdV binding: Coxsackie and Adenovirus Receptor, CD46, the glycans GD1a and polysialic acid, and desmoglein-2. Together with the integrin entry receptors, they display both symmetrical and asymmetrical modes of binding to the virus as demonstrated by the structural analyses reviewed here. The diversity of HAdV receptors contributes to the broad tropism of these viruses, and structural studies are thus an important source of information on HAdV-host cell interactions. The imbalance in structural data between the more and less extensively studied receptors remains to be addressed by future research.

9.
Viruses ; 11(10)2019 10 14.
Artigo em Inglês | MEDLINE | ID: mdl-31615155

RESUMO

Virus infections are initiated by the attachment of the viral particle to protein or carbohydrate receptors on the host cell. Sialic acid-bearing glycan structures are prominently displayed at the cell surface, and, consequently, these structures can function as receptors for a large number of diverse viruses. Structural biology research has helped to establish the molecular bases for many virus-sialic acid interactions. Due to the icosahedral 532 point group symmetry that underlies many viral capsids, the receptor binding sites are frequently arranged in a highly symmetric fashion and linked by five-fold, three-fold, or two-fold rotation axes. For the inhibition of viral attachment, one emerging strategy is based on developing multivalent sialic acid-based inhibitors that can simultaneously engage several of these binding sites, thus binding viral capsids with high avidity. In this review, we will evaluate the structures of non-enveloped virus capsid proteins bound to sialylated glycan receptors and discuss the potential of these structures for the development of potent antiviral attachment inhibitors.


Assuntos
Polissacarídeos/química , Ácidos Siálicos/química , Ligação Viral , Vírus/química , Antivirais/farmacologia , Sítios de Ligação , Cristalografia por Raios X , Modelos Moleculares , Receptores Virais , Vírus/efeitos dos fármacos
11.
Int J Med Microbiol ; 309(6): 151334, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31383542

RESUMO

The bacterial cell wall provides structural integrity to the cell and protects the cell from internal pressure and the external environment. During the course of the twelve-year funding period of the Collaborative Research Center 766, our work has focused on conducting structure-function studies of enzymes that modify (synthesize or cleave) cell wall components of a range of bacteria including Staphylococcus aureus, Staphylococcus epidermidis, and Nostoc punctiforme. Several of our structures represent promising targets for interference. In this review, we highlight a recent structure-function analysis of an enzyme complex that is responsible for the amidation of Lipid II, a peptidoglycan precursor, in S. aureus.


Assuntos
Proteínas de Bactérias/química , Proteínas de Bactérias/metabolismo , Complexos Multienzimáticos/química , Complexos Multienzimáticos/metabolismo , Peptidoglicano/metabolismo , Parede Celular/enzimologia , Parede Celular/metabolismo , Peptidoglicano/química , Domínios Proteicos , Staphylococcus/enzimologia , Staphylococcus/metabolismo , Relação Estrutura-Atividade , Uridina Difosfato Ácido N-Acetilmurâmico/análogos & derivados , Uridina Difosfato Ácido N-Acetilmurâmico/metabolismo
12.
Adv Carbohydr Chem Biochem ; 76: 65-111, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30851744

RESUMO

Sialic acid-based glycoconjugates cover the surfaces of many different cell types, defining key properties of the cell surface such as overall charge or likely interaction partners. Because of this prominence, sialic acids play prominent roles in mediating attachment and entry to viruses belonging to many different families. In this review, we first describe how interactions between viruses and sialic acid-based glycan structures can be identified and characterized using a range of techniques. We then highlight interactions between sialic acids and virus capsid proteins in four different viruses, and discuss what these interactions have taught us about sialic acid engagement and opportunities to interfere with binding.


Assuntos
Receptores Virais/química , Ácidos Siálicos/farmacologia , Viroses/tratamento farmacológico , Animais , Humanos , Receptores Virais/metabolismo , Ácidos Siálicos/química , Ácidos Siálicos/metabolismo , Viroses/virologia
13.
Nat Commun ; 10(1): 1320, 2019 03 21.
Artigo em Inglês | MEDLINE | ID: mdl-30899001

RESUMO

Attachment of human noroviruses to histo blood group antigens (HBGAs) is essential for infection, but how this binding event promotes the infection of host cells is unknown. Here, we employ protein NMR experiments supported by mass spectrometry and crystallography to study HBGA binding to the P-domain of a prevalent virus strain (GII.4). We report a highly selective transformation of asparagine 373, located in an antigenic loop adjoining the HBGA binding site, into an iso-aspartate residue. This spontaneous post-translational modification (PTM) proceeds with an estimated half-life of a few days at physiological temperatures, independent of the presence of HBGAs but dramatically affecting HBGA recognition. Sequence conservation and the surface-exposed position of this PTM suggest an important role in infection and immune recognition for many norovirus strains.


Assuntos
Asparagina/química , Antígenos de Grupos Sanguíneos/metabolismo , Proteínas do Capsídeo/química , Ácido Isoaspártico/química , Norovirus/metabolismo , Polissacarídeos/química , Processamento de Proteína Pós-Traducional , Asparagina/genética , Asparagina/metabolismo , Sítios de Ligação , Antígenos de Grupos Sanguíneos/química , Antígenos de Grupos Sanguíneos/genética , Proteínas do Capsídeo/genética , Proteínas do Capsídeo/metabolismo , Clonagem Molecular , Cristalografia por Raios X , Escherichia coli/genética , Escherichia coli/metabolismo , Expressão Gênica , Vetores Genéticos/química , Vetores Genéticos/metabolismo , Interações Hospedeiro-Patógeno , Humanos , Ácido Isoaspártico/genética , Ácido Isoaspártico/metabolismo , Cinética , Modelos Moleculares , Norovirus/genética , Polissacarídeos/metabolismo , Ligação Proteica , Conformação Proteica em alfa-Hélice , Conformação Proteica em Folha beta , Domínios e Motivos de Interação entre Proteínas , Multimerização Proteica , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo
14.
Macromol Biosci ; 19(5): e1800426, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30884172

RESUMO

Divalent precision glycooligomers terminating in N-acetylneuraminic acid (Neu5Ac) or 3'-sialyllactose (3'-SL) with varying linkers between scaffold and the glycan portions are synthesized via solid phase synthesis for co-crystallization studies with the sialic acid-binding major capsid protein VP1 of human Trichodysplasia spinulosa-associated Polyomavirus. High-resolution crystal structures of complexes demonstrate that the compounds bind to VP1 depending on the favorable combination of carbohydrate ligand and linker. It is found that artificial linkers can replace portions of natural carbohydrate linkers as long as they meet certain requirements such as size or flexibility to optimize contact area between ligand and receptor binding sites. The obtained results will influence the design of future high affinity ligands based on the structures presented here, and they can serve as a blueprint to develop multivalent glycooligomers as inhibitors of viral adhesion.


Assuntos
Proteínas do Capsídeo/química , Ácido N-Acetilneuramínico/química , Polyomavirus/química , Polissacarídeos/química , Cristalografia por Raios X , Humanos
15.
Sci Transl Med ; 10(470)2018 12 05.
Artigo em Inglês | MEDLINE | ID: mdl-30518612

RESUMO

Mutations in the gene encoding superoxide dismutase 1 (SOD1) lead to misfolding and aggregation of SOD1 and cause familial amyotrophic lateral sclerosis (FALS). However, the implications of wild-type SOD1 misfolding in sporadic forms of ALS (SALS) remain unclear. By screening human memory B cells from a large cohort of healthy elderly subjects, we generated a recombinant human monoclonal antibody (α-miSOD1) that selectively bound to misfolded SOD1, but not to physiological SOD1 dimers. On postmortem spinal cord sections from 121 patients with ALS, α-miSOD1 antibody identified misfolded SOD1 in a majority of cases, regardless of their SOD1 genotype. In contrast, the α-miSOD1 antibody did not bind to its epitope in most of the 41 postmortem spinal cord sections from non-neurological control (NNC) patients. In transgenic mice overexpressing disease-causing human SOD1G37R or SOD1G93A mutations, treatment with the α-miSOD1 antibody delayed the onset of motor symptoms, extended survival by up to 2 months, and reduced aggregation of misfolded SOD1 and motor neuron degeneration. These effects were obtained whether α-miSOD1 antibody treatment was administered by direct brain infusion or peripheral administration. These results support the further development of α-miSOD1 antibody as a candidate treatment for ALS involving misfolding of SOD1.


Assuntos
Esclerose Amiotrófica Lateral/tratamento farmacológico , Esclerose Amiotrófica Lateral/fisiopatologia , Anticorpos/uso terapêutico , Atividade Motora , Dobramento de Proteína/efeitos dos fármacos , Superóxido Dismutase-1/química , Superóxido Dismutase-1/metabolismo , Esclerose Amiotrófica Lateral/patologia , Animais , Anticorpos/administração & dosagem , Anticorpos/farmacologia , Modelos Animais de Doenças , Progressão da Doença , Humanos , Inflamação/patologia , Injeções Intraperitoneais , Injeções Intraventriculares , Camundongos Transgênicos , Atividade Motora/efeitos dos fármacos , Proteínas Recombinantes/farmacologia , Proteínas Recombinantes/uso terapêutico , Medula Espinal/metabolismo , Medula Espinal/patologia , Análise de Sobrevida
16.
Sci Rep ; 8(1): 18073, 2018 Dec 21.
Artigo em Inglês | MEDLINE | ID: mdl-30573753

RESUMO

A correction to this article has been published and is linked from the HTML and PDF versions of this paper. The error has not been fixed in the paper.

17.
Nature ; 563(7733): 705-709, 2018 11.
Artigo em Inglês | MEDLINE | ID: mdl-30464342

RESUMO

Methicillin-resistant Staphylococcus aureus (MRSA) is a frequent cause of difficult-to-treat, often fatal infections in humans1,2. Most humans have antibodies against S. aureus, but these are highly variable and often not protective in immunocompromised patients3. Previous vaccine development programs have not been successful4. A large percentage of human antibodies against S. aureus target wall teichoic acid (WTA), a ribitol-phosphate (RboP) surface polymer modified with N-acetylglucosamine (GlcNAc)5,6. It is currently unknown whether the immune evasion capacities of MRSA are due to variation of dominant surface epitopes such as those associated with WTA. Here we show that a considerable proportion of the prominent healthcare-associated and livestock-associated MRSA clones CC5 and CC398, respectively, contain prophages that encode an alternative WTA glycosyltransferase. This enzyme, TarP, transfers GlcNAc to a different hydroxyl group of the WTA RboP than the standard enzyme TarS7, with important consequences for immune recognition. TarP-glycosylated WTA elicits 7.5-40-fold lower levels of immunoglobulin G in mice than TarS-modified WTA. Consistent with this, human sera contained only low levels of antibodies against TarP-modified WTA. Notably, mice immunized with TarS-modified WTA were not protected against infection with tarP-expressing MRSA, indicating that TarP is crucial for the capacity of S. aureus to evade host defences. High-resolution structural analyses of TarP bound to WTA components and uridine diphosphate GlcNAc (UDP-GlcNAc) explain the mechanism of altered RboP glycosylation and form a template for targeted inhibition of TarP. Our study reveals an immune evasion strategy of S. aureus based on averting the immunogenicity of its dominant glycoantigen WTA. These results will help with the identification of invariant S. aureus vaccine antigens and may enable the development of TarP inhibitors as a new strategy for rendering MRSA susceptible to human host defences.


Assuntos
Parede Celular/química , Parede Celular/imunologia , Evasão da Resposta Imune , Staphylococcus aureus Resistente à Meticilina/citologia , Staphylococcus aureus Resistente à Meticilina/imunologia , Pentosefosfatos/imunologia , Ácidos Teicoicos/imunologia , Acetilglucosamina/química , Acetilglucosamina/metabolismo , Adulto , Animais , Bacteriófagos/patogenicidade , Feminino , Glicosilação , Glicosiltransferases/metabolismo , Humanos , Masculino , Staphylococcus aureus Resistente à Meticilina/química , Camundongos , Pessoa de Meia-Idade , Modelos Moleculares , Pentosefosfatos/química , Pentosefosfatos/metabolismo , Ácidos Teicoicos/química , Ácidos Teicoicos/metabolismo , Difosfato de Uridina/química , Difosfato de Uridina/metabolismo , Adulto Jovem
18.
Org Lett ; 20(22): 7201-7205, 2018 11 16.
Artigo em Inglês | MEDLINE | ID: mdl-30378433

RESUMO

Mutation of the gatekeeping residues for prenyl donor selectivity in six dimethylallyl transferases significantly increased their activities toward geranyl diphosphate. Forty-two geranylated derivatives were obtained from 15 cyclic dipeptides by using the engineered enzymes. Taking cyclo-l-Trp-l-Trp as an example, the geranyl moiety can be attached to all seven possible positions of the indole nucleus. This study demonstrates a convenient way to increase the structural diversity of geranylated products by structure-based engineering of the available dimethylallyl transferases.


Assuntos
Dimetilaliltranstransferase/química , Dipeptídeos/química , Difosfatos/química , Diterpenos/química , Alcaloides Indólicos/química , Peptídeos Cíclicos/química , Aspergillus/enzimologia , Clonagem Molecular , Dimetilaliltranstransferase/genética , Dipeptídeos/genética , Estrutura Molecular , Peptídeos Cíclicos/genética , Prenilação , Engenharia de Proteínas
19.
Viruses ; 10(11)2018 11 15.
Artigo em Inglês | MEDLINE | ID: mdl-30445709

RESUMO

Glycans are, with nucleic acids, proteins and lipids, one of the four founding structures of cellular life. Due to their non-template synthesis, they are inherently heterogeneous and difficult to study with regards to their structure and function. Since 2016, the research group ViroCarb, funded by the German Research Foundation, has investigated the role of glycans in non-enveloped virus infections with a highly interdisciplinary approach. The core idea was to bring together scientists and students from various disciplines such as structural biology, cell biology, virology and chemistry to advance research by an interdisciplinary means. In 2018, ViroCarb hosted the 1st International Symposium on Glycovirology in Schöntal, Germany, with a similar aim. Scientists from various disciplines gathered to discuss their area of study, present recent findings, establish or strengthen collaborations, and mentor the next generation of glycovirologists through formal presentations and informal discussions. The secluded meeting at the monastery of Schöntal gave ample time for in-depth discussions. On behalf of ViroCarb, this report summarizes the reports and highlights advances in the field.


Assuntos
Polissacarídeos/metabolismo , Viroses/patologia , Viroses/virologia , Vírus/química , Vírus/crescimento & desenvolvimento , Alemanha
20.
Org Biomol Chem ; 16(40): 7461-7469, 2018 10 17.
Artigo em Inglês | MEDLINE | ID: mdl-30270371

RESUMO

Microorganisms provide valuable enzyme machinery to assemble complex molecules. Fungal prenyltransferases (PTs) typically catalyse highly regiospecific prenylation reactions that are of significant pharmaceutical interest. While the majority of PTs accepts dimethylallyl diphosphate (DMAPP), very few such enzymes can use geranyl diphosphate (GPP) or farnesyl diphosphate (FPP) as donors. This catalytic gap prohibits the wide application of PTs for structural diversification. Structure-guided molecular modelling and site-directed mutagenesis of FgaPT2 from Aspergillus fumigatus led to the identification of the gatekeeping residue Met328 responsible for the prenyl selectivity and sets the basis for creation of GPP- and FPP-accepting enzymes. Site-saturation mutagenesis of the gatekeeping residue at position 328 in FgaPT2 revealed that the size of this side chain is the determining factor for prenyl selectivity, while its hydrophobicity is crucial for allowing DMAPP and GPP to bind.


Assuntos
Aspergillus fumigatus/enzimologia , Dimetilaliltranstransferase/química , Dimetilaliltranstransferase/genética , Engenharia de Proteínas , Prenilação de Proteína , Sequência de Aminoácidos , Dimetilaliltranstransferase/metabolismo , Modelos Moleculares , Conformação Proteica
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