Production of active recombinant eIF5A: reconstitution in E. coli of eukaryotic hypusine modification of eIF5A by its coexpression with modifying enzymes

Eukaryotic translation initiation factor 5A (eIF5A) is the only cellular protein that contains the polyamine-modified lysine, hypusine [Nε-(4-amino-2-hydroxybutyl)lysine]. Hypusine occurs only in eukaryotes and certain archaea, but not in eubacteria. It is formed post-translationally by two consecutive enzymatic reactions catalyzed by deoxyhypusine synthase (DHS) and deoxyhypusine hydroxylase (DOHH). Hypusine modification is essential for the activity of eIF5A and for eukaryotic cell proliferation. eIF5A binds to the ribosome and stimulates translation in a hypusine-dependent manner, but its mode of action in translation is not well understood. Since quantities of highly pure hypusine-modified eIF5A is desired for structural studies as well as for determination of its binding sites on the ribosome, we have used a polycistronic vector, pST39, to express eIF5A alone, or to co-express human eIF5A-1 with DHS or with both DHS and DOHH in Escherichia coli cells, to engineer recombinant proteins, unmodified eIF5A, deoxyhypusine- or hypusine-modified eIF5A. We have accomplished production of three different forms of recombinant eIF5A in high quantity and purity. The recombinant hypusine-modified eIF5A was as active in methionyl-puromycin synthesis as the native, eIF5A (hypusine form) purified from mammalian tissue. The recombinant eIF5A proteins will be useful tools in future structure/function and the mechanism studies in translation.

Expressão proteica das enzimas metabolizadoras de drogas no carcinoma de células escamosas de boca em pacientes jovens

Pós-graduação em Biopatologia Bucal - ICT

Estudo das alterações bioquímicas em sangue e diferentes órgãos de Phrynops geoffroanus (Schweigger, 1812) (Testudines: Chelidae) coletados em ambiente contaminado ou expostos ao benzo[a]pireno

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Chitin-degrading enzymes from an actinomycete ectosymbiont of Acromyrmex subterraneus brunneus (Hymenoptera: Formicidae)

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Pectinases From Sphenophorus levis Vaurie, 1978 (Coleoptera: Curculionidae): Putative Accessory Digestive Enzymes

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Marine-derived fungi: diversity of enzymes and biotechnological applications

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Piperine, a natural bioenhancer, nullifies the antidiabetic and antioxidant activities of curcumin in streptozotocin-diabetic rats

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

The multifaceted roles of metabolic enzymes in the Paracoccidioides species complex

Paracoccidioides species are dimorphic fungi and are the etiologic agents of paracoccidioidomycosis, which is a serious disease that involves multiple organs. The many tissues colonized by this fungus suggest a variety of surface molecules involved in adhesion. A surprising finding is that most enzymes in the glycolytic pathway, tricarboxylic acid (TCA) cycle and glyoxylate cycle in Paracoccidioides spp. have adhesive properties that aid in interacting with the host extracellular matrix and thus act as 'moonlighting'proteins. Moonlighting proteins have multiple functions, which adds a dimension to cellular complexity and benefit cells in several ways. This phenomenon occurs in both eukaryotes and prokaryotes. For example, moonlighting proteins from the glycolytic pathway or TCA cycle can play a role in bacterial pathogenesis by either acting as proteins secreted in a conventional pathway and/or as cell surface components that facilitate adhesion or adherence. This review outlines the multifunctionality exhibited by many Paracoccidioides spp. enzymes, including aconitase, aldolase, glyceraldehyde-3-phosphate dehydrogenase, isocitratelyase, malatesynthase, triose phosphate isomerase, fumarase, and enolase. We discuss the roles that moonlighting activities play in the virulence characteristics of this fungus and several other human pathogens during their interactions with the host.

Control of bacterial spot of tomato and activation of enzymes related to resistance by chemicals under field conditions

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Metabolismo do malondialdeído em peixes: implicações na avaliação da peroxidação lipídica como biomarcador de contaminação aquática

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)