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Under the guidance of genome mining combined with bioassay-coupled metabolomic analyses, an unprecedented macrodiolide sanyensin (1), with two flexible macrolides fused by the rigid oxabicyclo[3.3.1]nonane core, was isolated from the deep-sea-derived Streptomycessp. OUCT16-30. The stereochemistry of 1was established by NOEs (nuclear Overhauser effects), J-based configuration analysis, Marfey’s analysis, and together with a newly introduced stereochemical study workflow, which greatly shortens the time to obtain correct conformations of flexible structures based on the NMR constraints, thus leading to reliable quantum chemical calculations to establish the absolute configurations. This workflow is expected to have broad applicability for elucidating the stereochemistry of flexible natural products. The macrodiolide framework of 1is proposed to be formed through a biocatalytic cyclodimerization, followed by a series of nonenzymatic reactions. This work leads to new insights into the unexplored biosynthetic potential of deep-sea microbes and also provides a practical streamline for efficient mining of novel natural products, from discovery to stereochemical finalization.

Depression is immensely attributed to the overactivation of N-methyl-d-aspartic acid (NMDA) receptor in the brains. As regulatory binding partners of NMDA receptor, both Zn2+and H+are intimately interrelated to NMDA receptor’s activity. Therefore, exploring synergistic changes on the levels of Zn2+and H+in brains will promote the knowledge and treatment of depression. However, the lack of efficient, appropriate imaging tools limits simultaneously tracking Zn2+and H+in living mouse brains. Thus, a well-designed dual-color fluorescent probe (DNP) was fabricated for the simultaneous monitoring of Zn2+and H+in the brains of mice with depression. Encountering Zn2+, the probe evoked bright blue fluorescence at 460 nm. Meanwhile, the red fluorescence at 680 nm was decreased with H+addition. With blue/red dual fluorescence signal of DNP, we observed the synchronous increased Zn2+and H+in PC12 cells under oxidative stress. Notably, in vivoimaging for the first time revealed the simultaneous reduction of Zn2+and pH in brains of mice with depression-like behaviors. Further results implied that the NMDA receptor might be responsible for the coinstantaneous fluctuation of Zn2+and H+during depression. Altogether, this work is conducive to the knowledge of neural signal transduction mechanisms, advancing our understanding of the pathogenesis in depression.

Telomerase is a key enzyme for maintaining the telomere length and is regarded as a versatile cancer biomarker and a potential drug target due to its important role in cancer and aging. It is necessary to develop a sensitive and reliable method for detection of telomerase activity due to its very low level in cells. In this Article, we propose an ultrasensitive and robust method for quantitative determination of telomerase activity by combining single molecule fluorescence correlation spectroscopy (FCS) with telomerase repeat amplification protocol (TRAP). The principle of this new method (FCS-TRAP) is based on measurement of the change in characteristic diffusion time and molecule number of TRAP products by FCS. The characteristic diffusion time is related to the length of TRAP products, and the molecule number represents the concentration of TRAP products. We optimized the conditions of TRAP procedure and FCS measurements. We observed that the telomerase activities are positively correlated to characteristic diffusion time and molecule number of TRAP products at optimal conditions. This method was successfully used for determination of telomerase activity of different cells, and detection of a single cell was realized. Meanwhile, this method was used to evaluate the inhibition efficiency of inhibitors, and the IC50values obtained were in good agreement with the references. Compared to current TRAP methods, this method shows reliable quantification, ultrahigh sensitivity, and short detection time and is without separation. We believe that the FCS-TRAP method has a potential application in clinical diagnosis and screening of telomerase inhibitors.