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Characterization of butyrylcholinesterase from porcine milk.

ABSTRACT: Human butyrylcholinesterase (HuBChE) is under development for use as a pretreatment antidote against nerve agent toxicity. Animals are used to evaluate the efficacy of HuBChE for protection against organophosphorus nerve agents. Pharmacokinetic studies of HuBChE in minipigs showed a mean residence time of 267?h, similar to the half-life of HuBChE in humans, suggesting a high degree of similarity between BChE from 2 sources. Our aim was to compare the biochemical properties of PoBChE purified from porcine milk to HuBChE purified from human plasma. PoBChE hydrolyzed acetylthiocholine slightly faster than butyrylthiocholine, but was sensitive to BChE-specific inhibitors. PoBChE was 50-fold less sensitive to inhibition by DFP than HuBChE and 5-fold slower to reactivate in the presence of 2-PAM. The amino acid sequence of PoBChE determined by liquid chromatography tandem mass spectrometry was 91% identical to HuBChE. Monoclonal antibodies 11D8, mAb2, and 3E8 (HAH 002) recognized both PoBChE and HuBChE. Assembly of 4 identical subunits into tetramers occurred by noncovalent interaction with polyproline-rich peptides in PoBChE as well as in HuBChE, though the set of polyproline-rich peptides in milk-derived PoBChE was different from the set in plasma-derived HuBChE tetramers. It was concluded that the esterase isolated from porcine milk is PoBChE.

SUBMITTER: Saxena A

PROVIDER: S-EPMC6086383 | biostudies-literature | 2018 Aug

REPOSITORIES: biostudies-literature

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Characterization of butyrylcholinesterase from porcine milk.

Saxena Ashima A Belinskaya Tatyana T Schopfer Lawrence M LM Lockridge Oksana O

Archives of biochemistry and biophysics 20180615

Human butyrylcholinesterase (HuBChE) is under development for use as a pretreatment antidote against nerve agent toxicity. Animals are used to evaluate the efficacy of HuBChE for protection against organophosphorus nerve agents. Pharmacokinetic studies of HuBChE in minipigs showed a mean residence time of 267 h, similar to the half-life of HuBChE in humans, suggesting a high degree of similarity between BChE from 2 sources. Our aim was to compare the biochemical properties of PoBChE purified fro ...[more]

PMID: 29908755

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Project description:The composition of porcine milk oligosaccharides (PMO) was analyzed during early lactation and their relation to piglet gut microbiome was investigated. Pigs are considered ideal intestinal models to simulate humans because of the striking similarity in intestinal physiopathology to humans. The evolution of PMO was investigated in the milk from 3 healthy sows at prefarrowing, farrowing, and d 7 and 14 postpartum by Nano-LC Chip Quadrupole-Time-of-Flight mass spectrometer (Agilent Technologies, Santa Clara, CA). Previously sequenced metagenome libraries were reanalyzed to examine changes with specific gut bacterial populations. Over 30 oligosaccharides (OS) were identified in the milk, with 3'-sialyllactose, lacto-N-tetraose, ?1-3,?1-4-d-galactotriose, 2'-fucosyllactose, and 6'-sialyllactose being the most abundant species (accounting for ~70% of the total OS). Porcine milk had lower OS diversity (number of unique structures) than human milk, and appeared closer to bovine and caprine milk. In agreement with previous studies, only 3 fucosylated OS were identified. Surprisingly, their contribution to total OS abundance was greater than in bovine milk (9 vs. 1%). Indeed, fucosylated PMO increased during lactation, mirroring a similar trend observed for neutral and type I OS content during early lactation. Taken together, these results suggest that, in terms of abundance, PMO are closer to human milk than other domestic species, such as bovine and caprine milks. Metagenomic sequencing revealed that fucose-consuming bacterial taxa in the gut microbiota of piglets were qualitatively but not quantitatively different between nursing and weaning stages, suggesting that both the composition and structure of dietary glycans may play a critical role in shaping the distal gut microbiome. The similarity of both intestinal physiopathology and milk OS composition in human and porcine species suggests similar effects on gastrointestinal development of early nutrition, reinforcing the use of the pig intestinal model to simulate human intestinal models in the clinical setting.

Project description:BackgroundPorcine milk is a complex fluid, containing a myriad of immunological, biochemical, and cellular components, made to satisfy the nutritional requirements of the neonate. Whole milk contains many different cell types, including mammary epithelial cells, neutrophils, macrophages, and lymphocytes, as well nanoparticles, such as milk exosomes. To-date, only a limited number of livestock transcriptomic studies have reported sequencing of milk. Moreover, those studies focused only on sequencing somatic cells as a proxy for the mammary gland with the goal of investigating differences in the lactation process. Recent studies have indicated that RNA originating from multiple cell types present in milk can withstand harsh environments, such as the digestive system, and transmit regulatory molecules from maternal to neonate. Transcriptomic profiling of porcine whole milk, which is reflective of the combined cell populations, could help elucidate these mechanisms. To this end, total RNA from colostrum and mature milk samples were sequenced from 65 sows at differing parities. A stringent bioinformatic pipeline was used to identify and characterize 70,841 transcripts.ResultsThe 70,841 identified transcripts included 42,733 previously annotated transcripts and 28,108 novel transcripts. Differential gene expression analysis was conducted using a generalized linear model coupled with the Lancaster method for P-value aggregation across transcripts. In total, 1667 differentially expressed genes (DEG) were identified for the milk type main effect, and 33 DEG were identified for the milk type x parity interaction. Several gene ontology (GO) terms related to immune response were significant for the milk type main effect, supporting the well-known fact that immunoglobulins and immune cells are transferred to the neonate via colostrum.ConclusionsThis is the first study to perform global transcriptome analysis from whole milk samples in sows from different parities. Our results provide important information and insight into synthesis of milk proteins and innate immunity and potential targets for future improvement of swine lactation and piglet development.

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Characterization of butyrylcholinesterase from porcine milk.

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Characterization of butyrylcholinesterase from porcine milk.

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