Activity-based protein profiling (ABPP) has emerged as a powerful and versatile tool to enable annotation of protein functions and discovery of targets of bioactive ligands in complex biological systems. It utilizes chemical probes to covalently label functional sites in proteins so that they can be enriched for mass spectrometry (MS)-based quantitative proteomics analysis. However, the semistochastic nature of datadependent acquisition and high cost associated with isotopically encoded quantification reagents compromise the power of ABPP in multidimensional analysis and high-throughput screening, when a large number of samples need to be quantified in parallel. Here, we combine the data-independent acquisition (DIA) MS with ABPP to develop an efficient label-free quantitative chemical proteomic method, DIA-ABPP, with good reproducibility and high accuracy for high-throughput quantification. We demonstrated the power of DIA-ABPP for comprehensive profiling of functional cysteineome in three distinct applications, including dosedependent quantification of cysteines' sensitivity toward a reactive metabolite, screening of ligandable cysteines with a covalent fragment library, and profiling of cysteinome fluctuation in circadian clock cycles. DIA-ABPP will open new opportunities for indepth and multidimensional profiling of functional proteomes and interactions with bioactive small molecules in complex biological systems.

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Elucidating the isomeric structure of free fatty acids (FAs) in biological samples is essential to comprehend their biological functions in various physiological and pathological processes. Herein, we report a novel...

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To investigate key taste-active components in Takifugu obscurus (T. obscurus), twenty-eight putative taste compounds in cooked muscle of T. obscurus were quantitatively analyzed and the pivotal components were identified by taste reconstitution, omission and addition tests. Moreover, the role of flavor peptides in overall taste profile of T. obscurus were evaluated. Sensory evaluation revealed that glutamic acid, serine, proline, arginine, lysine, adenosine-5´-monophosphate, inosine-5´-monophosphate (IMP), succinic acid, sodium, potassium, phosphates and chlorides, were the core taste-active contributors to T. obscurus. Besides glutamic acid, IMP, succinic acid and potassium, the characteristic T. obscurus-like umami and kokumi profiles were induced by adding flavor peptides, among which Pro-Val-Ala-Arg-Met-Cys-Arg and Tyr-Gly-Gly-Thr-Pro-Pro-Phe-Val were identified as key substances on the basis of addition test and dose-response analysis.The present data may help to reveal the secret of delicious taste of T. obscurus and provide the basis for development of deeper flavor analysis of pufferfish.

Abstract:A high affinity polyclonal antibody-based enzyme linked immunosorbent assay (ELISA) was developed for the quantification of zeranol in bovine urine. On the basis of urine matrix studies, the optimized dilution factors producing insignificant matrix interference were selected as 1:5 in pretreatment. In the improved ELISA, the linear response range was between 0.02 and 1 µg/ml , and the detection limit was 0.02 µg/ml for the assay. The overall recoveries and the coefficients of variation (CVs) were in the range of 82%~127% and 3.5%~8.8%, respectively. Thirty-six bovine urine samples spiked with zeranol (ranging from 0.2 to 10 µg/ml) were detected by the ELISA and liquid chromatography (LC) method, and good correlations were obtained between the two methods (R 2 =0.9643). We conclude that this improved ELISA is suitable tool for a mass zeranol screening and can be an alternative for the conventional LC method for zeranol in bovine urine.

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