Summary Bacterial aromatic polyketides are a group of natural products synthesized by polyketide synthases (PKSs) that show diverse structures and biological activities. They are structurally sub-classified into linear, angular, and discoid aromatic polyketides, the formation of which is commonly determined by the shaping and folding of the poly-β-keto intermediates under the concerted actions of the minimal PKSs, cyclases and ketoreductases. Murayaquinone, found in several streptomycetes, possesses an unusual tricyclic angular aromatic polyketide core containing a 9,10-phenanthraquinone. In this study, genes essential for murayaquinone biosynthesis were identified, and a linear anthraoxirene intermediate was discovered. A unique biosynthetic model for the angular aromatic polyketide formation was discovered and confirmed through in vivo and in vitro studies. Three oxidoreductases, MrqO3, MrqO6, and MrqO7, were identified to catalyze the conversion of the linear aromatic polyketide intermediate into the final angularly arranged framework, which exemplifies a novel strategy for the biosynthesis of angular aromatic polyketides.

c Lipstatin, isolated from Streptomyces toxytricini as a potent and selective inhibitor of human pancreatic lipase, is a precursor for tetrahydrolipstatin (also known as orlistat, Xenical, and Alli), the only FDA-approved antiobesity medication for long-term use. Lipstatin features a 2-hexyl-3,5-dihydroxy-7,10-hexadecadienoic-␤-lactone structure with an N-formyl-L-leucine group attached as an ester to the 5-hydroxy group. It has been suggested that the ␣-branched 3,5-dihydroxy fatty acid ␤-lactone moiety of lipstatin in S. toxytricini is derived from Claisen condensation between two fatty acid substrates, which are derived from incomplete oxidative degradation of linoleic acid based on feeding experiments. In this study, we identified a six-gene operon (lst) that was essential for the biosynthesis of lipstatin by large-deletion, complementation, and single-gene knockout experiments. lstA, lstB, and lstC, which encode two ␤-ketoacyl-acyl carrier protein synthase III homologues and an acyl coenzyme A (acyl-CoA) synthetase homologue, were indicated to be responsible for the generation of the ␣-branched 3,5-dihydroxy fatty acid backbone. Subsequently, the nonribosomal peptide synthetase (NRPS) gene lstE and the putative formyltransferase gene lstF were involved in decoration of the ␣-branched 3,5-dihydroxy fatty acid chain with an N-formylated leucine residue. Finally, the 3␤-hydroxysteroid dehydrogenase-homologous gene lstD might be responsible for the reduction of the ␤-keto group of the biosynthetic intermediate, thereby facilitating the formation of the unique ␤-lactone ring.

Streptomyces lividans is a suitable host for the heterologous expression of biosynthetic gene clusters (BGCs) from actinomycetes to discover “cryptic” secondary metabolites. To improve the heterologous expression of BGCs, herein we optimized S. lividans strain SBT5 via the stepwise integration of three global regulatory genes and two codon-optimized multi-drug efflux pump genes and deletion of a negative regulatory gene, yielding four engineered strains. All optimization steps were observed to promote the heterologous production of polyketides, non-ribosomal peptides, and hybrid antibiotics. The production increments of these optimization steps were additional, so that the antibiotic yields were several times or even dozens of times higher than the parent strain SBT5 when the final optimized strain, S. lividans LJ1018, was used as the heterologous expression host. The heterologous production of these antibiotics in S. lividans LJ1018 and GX28 was also much higher than in the strains from which the BGCs were isolated. S. lividans LJ1018 and GX28 markedly promoted the heterologous production of secondary metabolites, without requiring manipulation of gene expression components such as promoters on individual gene clusters. Therefore, these strains are well-suited as heterologous expression hosts for secondary metabolic BGCs. In addition, we successfully conducted high-throughput library expression and functional screening (LEXAS) of one bacterial artificial chromosome library and two cosmid libraries of three Streptomyces genomes using S. lividans GX28 as the library-expression host. The LEXAS experiments identified clones carrying intact BGCs sufficient for the heterologous production of piericidin A1, murayaquinone, actinomycin D, and dehydrorabelomycin. Notably, due to lower antibiotic production, the piericidin A1 BGC had been overlooked in a previous LEXAS screening using S. lividans SBT5 as the expression host. These results demonstrate the feasibility and superiority of S. lividans GX28 as a host for high-throughput screening of genomic libraries to mine cryptic BGCs and bioactive compounds.

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Nerve growth factor (NGF) is important for peripheral nerve regeneration. However, its short half‐life and rapid diffusion in body fluids limit its clinical efficacy. Collagen has favorable biocompatibility and biodegradability, and weak immunogenicity. Because it possesses an NGF binding domain, we cross‐linked heparin to collagen tubes to construct nerve guidance conduits for delivering NGF. The conduits were implanted to bridge a facial nerve defect in rats. Histological and functional analyses were performed to assess the effect of the nerve guidance conduit on facial nerve regeneration. Heparin enhanced the binding of NGF to collagen while retaining its bioactivity. Also, the nerve guidance conduit significantly promoted axonal growth and Schwan cell proliferation at 12 weeks after surgery. The nerve regeneration and functional recovery outcomes using the nerve guidance conduit were similar to those of autologous nerve grafting. Therefore, the nerve guidance conduit may promote safer nerve regeneration.