Discovery and Characterization of a Myxobacterial Lanthipeptide with Unique Biosynthetic Features and Anti-inflammatory Activity.

The genomes of myxobacteria harbor a variety of biosynthetic gene clusters encoding numerous secondary metabolites, including ribosomally synthesized and post-translationally modified peptides (RiPPs) with diverse chemical structures and biological activities. However, the biosynthetic potential of RiPPs from myxobacteria remains barely explored. Herein, we report a novel myxobacteria lanthipeptide myxococin identified from Myxococcus fulvus. Myxococins represent the first example of lanthipeptides, of which the characteristic multiple thioether rings are installed by employing a Class II lanthipeptide synthetase MfuM and a Class I lanthipeptide cyclase MfuC in a cascaded way. Unprecedentedly, we biochemically characterized the first M61 family aminopeptidase MfuP involved in RiPP biosynthesis, demonstrating that MfuP showed the activity of an endopeptidase activity. MfuP is leader-independent but strictly selective for the multibridge structure of myxococin A and responsible for unwrapping two rings via amide bond hydrolysis, yielding myxococin B. Furthermore, the X-ray crystal structure of MfuP and structural analysis, including active-site mutations, are reported. Finally, myxococins are evaluated to exhibit anti-inflammatory activity in lipopolysaccharide-induced macrophages without detectable cytotoxicity.

Recent Advances in Discovery, Bioengineering, and Bioactivity-Evaluation of Ribosomally Synthesized and Post-translationally Modified Peptides.

Ribosomally synthesized and post-translationally modified peptides (RiPPs) are of increasing interest in natural products as well as drug discovery. This empowers not only the unique chemical structures and topologies in natural products but also the excellent bioactivities such as antibacteria, antifungi, antiviruses, and so on. Advances in genomics, bioinformatics, and chemical analytics have promoted the exponential increase of RiPPs as well as the evaluation of biological activities thereof. Furthermore, benefiting from their relatively simple and conserved biosynthetic logic, RiPPs are prone to be engineered to obtain diverse analogues that exhibit distinct physiological activities and are difficult to synthesize. This Review aims to systematically address the variety of biological activities and/or the mode of mechanisms of novel RiPPs discovered in the past decade, albeit the characteristics of selective structures and biosynthetic mechanisms are briefly covered as well. Almost one-half of the cases are involved in anti-Gram-positive bacteria. Meanwhile, an increasing number of RiPPs related to anti-Gram-negative bacteria, antitumor, antivirus, etc., are also discussed in detail. Last but not least, we sum up some disciplines of the RiPPs' biological activities to guide genome mining as well as drug discovery and optimization in the future.

Engineered Biosynthesis of Complex Disorazol Polyketides in a Streamlined Burkholderia thailandensis.

Engineering the biosynthetic pathways of complex natural products is a significant approach to obtain derivatives with improved properties. Here, we constructed a streamlined engineered biosynthesis system of myxobacterium-derived complex polyketide disorazol in a heterologous host, Burkholderia thailandensis E264. Inactivation of dehydratase domains in the disorazol biosynthetic pathway led to the production of two hydroxylated derivatives. Module deletion allowed the generation of an unnatural derivative with a truncated macrolactone ring, and the ACP-KS linker was the optimal fusion region for module deletion in this trans-AT polyketide synthase. These disorazol derivatives showed different activities against human cancer cell lines ranging from the nanomolar to micromolar level, suggesting the primary structure-activity relationship. The PKS engineering enables structural derivatization of disorazol, facilitating the in-depth engineered biosynthesis of polyketides.

Engineering of Burkholderia thailandensis strain E264 serves as a chassis for expression of complex specialized metabolites.

Heterologous expression is an indispensable approach to exploiting natural products from phylogenetically diverse microbial communities. In this study, we constructed a heterologous expression system based on strain Burkholderia thailandensis E264 by deleting efflux pump genes and screening constitutive strong promoters. The biosynthetic gene cluster (BGC) of disorazol from Sorangium cellulosum So ce12 was expressed successfully with this host, and the yield of its product, disorazol F2, rather than A1, was improved to 38.3 mg/L by promoter substitution and insertion. In addition to the disorazol gene cluster, the BGC of rhizoxin from Burkholderia rhizoxinica was also expressed efficiently, whereas no specific peak was detected when shuangdaolide BGC from Streptomyces sp. B59 was transformed into the host. This system provides another option to explore natural products from different phylogenetic taxa.

Fulvivirga marina sp. nov. and Fulvivirga sediminis sp. nov., two novel Bacteroidetes isolated from the marine sediment.

Two novel, designated strains 29W222T and 2943T, were isolated from the marine sediment from Aoshan Bay, Jimo, PR China. Growth was observed at pH 6.0-8.5 (optimum, pH 7.5) for strain 29W222T, and pH 5.5-8.5 (pH 7.0) for strain 2943T. Both strains displayed growth in 0.5-6 % NaCl with an optimum at 1 % for 29W222T; 0.5 % for 2943T. Both strains grew optimally at 33 °C. The results of phylogenetic analyses based on 16S rRNA gene sequences indicated that 29W222T and 2943T represented members of the genus Fulvivirga and strain 29W222T was most closely related to Fulvivirga kasyanovii KMM 6220T (97.9 % sequence similarity) and Fulvivirga imtechensis AK7T (95.0 %), and 2943T to Fulvivirga imtechensis AK7T (95.7 %) and Fulvivirga kasyanovii KMM 6220T (94.8 %). The genomic DNA G+C contents of 29W222T and 2943T were 39.9 and 37.7 mol%, respectively. The results of chemotaxonomic analysis indicated that the sole respiratory quinone was menaquinone 7 (MK-7), and the major fatty acid was iso-C15 : 0 for both strains. Average nucleotide identity and average amino acid identity values between strain 29W222T and Fulvivirga kasyanovii KMM 6220T were 78.9 and 83.6 %, respectively; the corresponding values between 2943T and Fulvivirga imtechensis AK7T were 69.8 and 63.6 %, respectively. Therefore, strains 29W222T and 2943T represent to two novel species of the genus Fulvivirga, for which the names Fulvivirga marina sp. nov. (29W222T=KCTC 62848T=MCCC 1K05194T) and Fulvivirga sediminis sp. nov. (2943T=KCTC 62847T= MCCC 1K05144T) are proposed, respectively.

Genome Mining and Biosynthesis of Primary Amine-Acylated Desferrioxamines in a Marine Gliding Bacterium.

Genome mining of Fulvivirga sp. W222 revealed a desferrioxamine-like biosynthetic gene cluster containing an unknown gene fulF that is conserved in many Bacteroidetes species. A series of primary amine-acylated desferrioxamine G1 analogues, fulvivirgamides, were identified, and fulvivirgamides A2, B2, B3, and B4 (1-4) were purified and characterized. The function of FulF, which is a novel acyltransferase for the acylation of the primary amine of Desferrioxamine G1, was verified by heterologous expression and feeding experiments.

Algoriphagus resistens sp. nov., isolated from marine sediment.

Strain NH1T, a pink-pigmented, facultatively anaerobic, heterotrophic, catalase-positive and oxidase-negative, Gram-stain-negative marine bacterium, was isolated from marine sediment on the coast of Weihai, China. Cells of strain NH1T were rod-shaped, 0.8-2.0 µm in length and 0.5-1.0 µm in width. The strain was able to grow at 13-37 °C, pH 5.5-8.5, in the presence of 0.0-8.0 % (w/v) NaCl. Optimal growth was observed at 28 °C, with 3.0 % (w/v) NaCl and pH 6.5-7.0. Nitrate was reduced. The G+C content of the DNA was 41.9 mol%. The major isoprenoid quinone was MK-7 and the main cellular fatty acids (>10 %) were summed feature 3 (33.6 %) comprising iso-C15 : 0 2-OH and/or C16 : 1ω7c, and iso-C15:0 (19.2%). The major polar lipids in strain NH1T were phosphatidylethanolamine, unidentified lipids, phospholipid and aminolipids. Phylogenetic analysis based on 16S rRNA gene sequences revealed that strain NH1T was highly related to the type strains of Algoriphagus antarcticus (97.87 % 16SrRNA gene sequence similarity) and Algoriphagus ratkowskyi (97.56 %). On basis of the phenotypic and phylogenetic data, strain NH1T should be classified as representing a novel species of the genus Algoriphagus, for which the name Algoriphagus resistens sp. nov. is proposed. The type strain is NH1T (=MCCC 1H00140T=KCTC 52228T).

Woeseia oceani gen. nov., sp. nov., a chemoheterotrophic member of the order Chromatiales, and proposal of Woeseiaceae fam. nov.

A novel Gram-stain-negative, rods or bent rods, facultatively anaerobic, oxidase-negative and catalase-positive bacterium, designated XK5T, was isolated from coastal sediment from Xiaoshi Island, Weihai, China. Optimal growth occurred at 28-35 °C (range 8-42 °C) and pH 7.0-8.0 (range pH 6.0-9.0) with 1-3 % (w/v) NaCl (range 0.5-8 %). Phylogenetic analysis based on 16S rRNA gene sequences revealed that strain XK5T was 92.1 % similar to the type strain of Thioalkalivibrio thiocyanodenitrificans, 91.9 % to the type strain of Thioalkalivibrio sulfidiphilus and 91.8 % to the type strain of Thioalkalivibrio denitrificans; similarity to other species was less than 91 %. The isolate and closely related environmental clones formed a novel family level clade in the order Chromatiales. The polar lipid profile of the novel isolate consisted of phosphatidylethanolamine, phosphatidylglycerol and some other unknown phospholipids, aminolipids and lipids. Major cellular fatty acids were iso-C17 : 1ω9c and iso-C15 : 0 and the main respiratory lipoquinone was Q-8. The DNA G+C content of strain XK5T was 59.3 mol%. Comparative analysis of 16S rRNA gene sequences and characterization indicated that strain XK5T represents a novel species of a new genus within a novel family of the order Chromatiales, for which the name Woeseia oceani gen. nov., sp. nov. is proposed. The type strain of Woeseia oceani is XK5T ( = ATCC BAA-2615T = CICC 10905T). In addition, a novel family name, Woeseiaceae fam. nov., is proposed to accommodate the genus Woeseia.

Bradymonas sediminis gen. nov., sp. nov., isolated from coastal sediment, and description of Bradymonadaceae fam. nov. and Bradymonadales ord. nov.

A novel Gram-stain-negative, rod-shaped, gliding, facultatively anaerobic, oxidase-negative and catalase-positive bacterium, designated FA350(T), was isolated from coastal sediment from Xiaoshi Island, Weihai, China. Strain FA350(T) showed growth on modified nutrient agar supplemented with 0.1% d-(+)-trehalose and with distilled water replaced by seawater. Optimal growth occurred at 33 °C and pH 8.5 with 4% NaCl. Phylogenetic analysis based on 16S rRNA gene sequences revealed that strain FA350(T) belongs to a novel bacterial order in the class Deltaproteobacteria , and the most closely related type strains belong to the order Desulfuromonadales , with 85.1-85.6% 16S rRNA gene sequence similarity. The polar lipid profile of the novel strain consisted of phosphatidylethanolamine, phosphatidylglycerol and two unknown phospholipids. Major cellular fatty acids were iso-C15 : 0, iso-C17 : 0 and iso-C17 : 1ω10c and menaquinone MK-7 was the sole respiratory quinone. The DNA G+C content of strain FA350(T) was 60.3 mol%. The isolate and closely related environmental clones formed a novel order-level clade in the class Deltaproteobacteria . Comparative analysis of 16S rRNA gene sequences and characterization indicated that strain FA350(T) may represent a novel order of the Deltaproteobacteria . Here, we propose the name Bradymonas sediminis gen. nov., sp. nov. to accommodate strain FA350(T). The type strain of Bradymonas sediminis is FA350(T) ( =DSM 28820(T) =CICC 10904(T)); Bradymonadales ord. nov. and Bradymonadaceae fam. nov. are also proposed to accommodate the novel taxon.

Motiliproteus sediminis gen. nov., sp. nov., isolated from coastal sediment.

A novel Gram-stain-negative, rod-to-spiral-shaped, oxidase- and catalase- positive and facultatively aerobic bacterium, designated HS6(T), was isolated from marine sediment of Yellow Sea, China. It can reduce nitrate to nitrite and grow well in marine broth 2216 (MB, Hope Biol-Technology Co., Ltd) with an optimal temperature for growth of 30-33 °C (range 12-45 °C) and in the presence of 2-3% (w/v) NaCl (range 0.5-7%, w/v). The pH range for growth was pH 6.2-9.0, with an optimum at 6.5-7.0. Phylogenetic analysis based on 16S rRNA gene sequences demonstrated that the novel isolate was 93.3% similar to the type strain of Neptunomonas antarctica, 93.2% to Neptunomonas japonicum and 93.1% to Marinobacterium rhizophilum, the closest cultivated relatives. The polar lipid profile of the novel strain consisted of phosphatidylethanolamine, phosphatidylglycerol and some other unknown lipids. Major cellular fatty acids were summed feature 3 (C(16:1) ω7c/iso-C15:0 2-OH), C(18:1) ω7c and C(16:0) and the main respiratory quinone was Q-8. The DNA G+C content of strain HS6(T) was 61.2 mol%. Based on the phylogenetic, physiological and biochemical characteristics, strain HS6(T) represents a novel genus and species and the name Motiliproteus sediminis gen. nov., sp. nov., is proposed. The type strain is HS6(T) (=ATCC BAA-2613(T)=CICC 10858(T)).