Discovery of Terminal Oxazole-Bearing Natural Products by a Targeted Metabologenomic Approach.

A targeted metabologenomic method was developed to selectively discover terminal oxazole-bearing natural products from bacteria. For this, genes encoding oxazole cyclase, a key enzyme in terminal oxazole biosynthesis, were chosen as the genomic signature to screen bacterial strains that may produce oxazole-bearing compounds. Sixteen strains were identified from the screening of a bacterial DNA library (1,000 strains) using oxazole cyclase gene-targeting polymerase chain reaction (PCR) primers. The PCR amplicon sequences were subjected to phylogenetic analysis and classified into nine clades. 1H-13C coupled-HSQC NMR spectra obtained from the culture extracts of the hit strains enabled the unequivocal detection of the target compounds, including five new oxazole compounds, based on the unique 1JCH values and chemical shifts of oxazole: lenzioxazole (1) possessing an unprecedented cyclopentane, permafroxazole (2) bearing a tetraene conjugated with carboxylic acid, tenebriazine (3) incorporating two modified amino acids, and methyl-oxazolomycins A and B (4 and 5). Tenebriazine displayed inhibitory activity against pathogenic fungi, whereas methyl-oxazolomycins A and B (4 and 5) selectively showed anti-proliferative activity against estrogen receptor-positive breast cancer cells. This metabologenomic method enables the logical and efficient discovery of new microbial natural products with a target structural motif without the need for isotopic labeling.

Taeanamides A and B, Nonribosomal Lipo-Decapeptides Isolated from an Intertidal-Mudflat-Derived Streptomyces sp.

Two new lipo-decapeptides, namely taeanamides A and B (1 and 2), were discovered from the Gram-positive bacterium Streptomyces sp. AMD43, which was isolated from a mudflat sample from Anmyeondo, Korea. The exact molecular masses of 1 and 2 were revealed by high-resolution mass spectrometry, and the planar structures of 1 and 2 were elucidated using NMR spectroscopy. The absolute configurations of 1 and 2 were determined using a combined analysis of 1H-1H coupling constants and ROESY correlations, the advanced Marfey's method, and bioinformatics. The putative nonribosomal peptide synthetase pathway for the taeanamides was identified by analyzing the full genome sequence data of Streptomyces sp. AMD43. We also found that taeanamide A exhibited mild anti-tuberculosis bioactivity, whereas taeanamide B showed significant bioactivity against several cancer cell lines.

Gwanakosides A and B, 6-Deoxy-α-l-talopyranose-Bearing Aromatic Metabolites from a Streptomyces sp. and Coculture with Pandoraea sp.

Single-strain cultivation of a mountain soil-derived Streptomyces sp. GA02 and its coculture with Pandoraea sp. GA02N produced two aromatic products, gwanakosides A and B (1 and 2, respectively). Their spectroscopic analysis revealed that 1 is a new dichlorinated naphthalene glycoside and 2 is a pentacyclic aromatic glycoside. The assignment of the two chlorine atoms in 1 was confirmed by the analysis of its band-selective CLIP-HSQMBC spectrum. The sugars in the gwanakosides were identified as 6-deoxy-α-l-talopyranose based on 1H-1H coupling constants, Rotating frame Overhauser enhancement spectroscopy (ROESY) NMR correlations, and chemical derivatization followed by spectroscopic and chromatographic analyses. The absolute configuration of 2, whose production was enhanced approximately 100-fold in coculture, was proposed based on a quantum mechanics-based chemical shift analysis method, DP4 calculations, and the chemically determined configuration of 6-deoxy-α-l-talopyranose. Gwanakoside A displayed inhibitory activity against pathogenic bacteria, including Staphylococcus aureus (MIC = 8 µg/mL) and Mycobacterium tuberculosis (MIC50 = 15 µg/mL), and antiproliferative activity against several human cancer cell lines (IC50 = 5.6-19.4 µM).

Effects of the Crack Tip Constraint on the Fracture Assessment of an Al 5083-O Weldment for Low Temperature Applications.

The constraint effect is the key issue in structural integrity assessments based on two parameter fracture mechanics (TPFM) to make a precise prediction of the load-bearing capacity of cracked structural components. In this study, a constraint-based failure assessment diagram (FAD) was used to assess the fracture behavior of an Al 5083-O weldment with various flaws at cryogenic temperature. The results were compared with those of BS 7910 Option 1 FAD, in terms of the maximum allowable stress. A series of fracture toughness tests were conducted with compact tension (CT) specimens at room and cryogenic temperatures. The Q parameter for the Al 5083-O weldment was evaluated to quantify the constraint level, which is the difference between the actual stress, and the Hutchinson-Rice-Rosengren (HRR) stress field near the crack tip. Nonlinear 3D finite element analysis was carried out to calculate the Q parameter at cryogenic temperature. Based on the experimental and numerical results, the influence of the constraint level correction on the allowable applied stress was investigated using a FAD methodology. The results showed that the constraint-based FAD procedure is essential to avoid an overly conservative allowable stress prediction in an Al 5083-O weldment with flaws.