RESUMO
A putative Type III Polyketide synthase (PKSIII) encoding gene was identified from a marine yeast, Naganishia uzbekistanensis strain Mo29 (UBOCC-A-208024) (formerly named as Cryptococcus sp.) isolated from deep-sea hydrothermal vents. This gene is part of a distinct phylogenetic branch compared to all known terrestrial fungal sequences. This new gene encodes a C-terminus extension of 74 amino acids compared to other known PKSIII proteins like Neurospora crassa. Full-length and reduced versions of this PKSIII were successfully cloned and overexpressed in a bacterial host, Escherichia coli BL21 (DE3). Both proteins showed the same activity, suggesting that additional amino acid residues at the C-terminus are probably not required for biochemical functions. We demonstrated by LC-ESI-MS/MS that these two recombinant PKSIII proteins could only produce tri- and tetraketide pyrones and alkylresorcinols using only long fatty acid chain from C8 to C16 acyl-CoAs as starter units, in presence of malonyl-CoA. In addition, we showed that some of these molecules exhibit cytotoxic activities against several cancer cell lines.
Assuntos
Antineoplásicos/metabolismo , Basidiomycota/enzimologia , Proteínas Fúngicas/metabolismo , Policetídeo Sintases/metabolismo , Policetídeos/metabolismo , Antineoplásicos/farmacologia , Células CACO-2 , Sobrevivência Celular/efeitos dos fármacos , Proteínas Fúngicas/isolamento & purificação , Proteínas Fúngicas/farmacologia , Humanos , Fontes Hidrotermais/microbiologia , Neoplasias/tratamento farmacológico , Neoplasias/patologia , Filogenia , Policetídeo Sintases/isolamento & purificação , Policetídeo Sintases/farmacologia , Policetídeos/farmacologia , Especificidade por Substrato , Células THP-1 , Microbiologia da ÁguaRESUMO
Because of their ecological functions, natural products have been optimized in evolution for interaction with biological systems and receptors. However, they have not necessarily been optimized for other desirable drug properties and thus can often be improved by structural modification. Using examples from the literature, this paper reviews the opportunities for increasing structural diversity among natural products by combinatorial biosynthesis, i.e., the genetic manipulation of biosynthetic pathways. It distinguishes between combinatorial biosynthesis in a narrower sense to generate libraries of modified structures, and metabolic engineering for the targeted formation of specific structural analogs. Some of the problems and limitations encountered with these approaches are also discussed. Work from the author's laboratory on ansamycin antibiotics is presented which illustrates some of the opportunities and limitations.