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Gao C.,XingHuaLing District Food and Drug Administration | Jin M.,Third Institute of Oceanography | Yi Z.,Third Institute of Oceanography | Zeng R.,Third Institute of Oceanography | And 2 more authors.
Journal of Microbiology and Biotechnology | Year: 2015

A novel sulfatase gene, ary423 (1,536 bp ORF), encoding a protein of 511 amino acids with a calculated molecular mass of 56 kDa, was identified from Flammeovirga pacifica, which was isolated from deep-sea sediments of west Pacific Ocean. Amino acid sequence analysis revealed that Ary423 possessed a conserved C-X-A-X-R motif, which was recognized as the sulfatase signature. Phylogenetic analysis suggested that Ary423 belonged to arylsulfatases. After heterologous expression in Escherichia coli cells, the recombinant Ary423 was purified with a Ni+ affinity column, and was shown to be highly active at a broad range of temperatures from 30° to 70°C, with maximum activity at 40°C. Furthermore, recombinant Ary423 retained more than 70% and 40% of its maximum activity after 12 h of incubation at 50°C and 60°C, respectively, exhibiting good thermostability at high temperatures. The optimal pH for Ary423 was determined to be 8.0 and the activity of Ary423 could be slightly enhanced by Mg2+. The recombinant enzyme could hydrolyze sulfate ester bonds in p-nitrophenyl sulfate (NPS) and Asparagus crude polysaccharides with a specific activity of 64.8 U/mg and 25.4 U/mg, respectively. These favorable properties could make Ary423 attractive for application in the desulfating process of agar production. © 2015 by The Korean Society for Microbiology and Biotechnology. Source


Shu L.,Xiamen University | Yang Y.,Xiamen University | Huang H.,Xiamen University | Ye H.,Xiamen University | Ye H.,Fujian Collaborative Innovation Center for Exploitation and Utilization of Marine Biological Resources
Molecular and Cellular Endocrinology | Year: 2016

In vertebrates, bone morphogenetic proteins (BMPs) play an important role in various biological processes. However, the function of BMPs in crustaceans is still unknown. In our study, a ligand (BMP7) and two receptors (Sp-BMPRIB and Sp-BMPRII) are cloned firstly in the mud crab, Scylla paramamosain. The qRT-PCR demonstrated that both ligand and receptors were expressed in various tissues, especially in ovary. The expression of BMPRs mRNA increased along the ovarian development, while BMP7 had an opposite tendency. In-situ hybridization revealed that Sp-BMPRIB and Sp-BMPRII were expressed in both oocytes and follicle cells, whereas Sp-BMP7 was exclusively localized in follicle cells. RNAi experiments showed that the expression levels of Smad1 and vitellogenin receptor declined rapidly after BMPRs were silenced. Based on these data, we hypothesized that in S. paramamosain, BMP7 and BMPRs had impact on the ovarian development, presumably via the autocrine/paracrine way. © 2016 Elsevier Ireland Ltd Source


Zhang Y.,State Oceanic Administration | Zhang Y.,Fujian Collaborative Innovation Center for Exploitation and Utilization of Marine Biological Resources | Chen D.,China National Center for Food Safety Risk Assessment | Hong Z.,State Oceanic Administration | Hong Z.,China National Center for Food Safety Risk Assessment
Toxins | Year: 2015

In this study, we developed a self-assembly pipette tip solid-phase extraction (PTSPE) method using a high molecular weight polymer material (PAX) as the adsorbent for the determination of domoic acid (DA) in human urine samples by liquid chromatography high-resolution mass spectrometry (LC-HRMS) analysis. The PTSPE cartridge, assembled by packing 9.1 mg of PAX as sorbent into a 200 μL pipette tip, showed high adsorption capacity for DA owing to the strong cationic properties of PAX. Compared with conventional SPE, the PTSPE is simple and fast, and shows some advantages in the aspects of less solvent consumption, low cost, the absence of the evaporation step, and short time requirement. All the parameters influencing the extraction efficiency such as pH, the amount of sorbent, the number of aspirating/dispensing cycles, and the type and volume of eluent in PTSPE were carefully investigated and optimized. Under the optimized conditions, the limit of detection (LOD) and limit of quantification (LOQ) values of DA were 0.12 μg/L and 0.37 μg/L respectively. The extraction recoveries of DA from the urine samples spiked at four different concentrations were in a range from 88.4% to 102.5%. The intra- and inter-day precisions varied from 2.1% to 7.6% and from 2.6% to 12.7%, respectively. The accuracy ranged from -1.9% to -7.4%. © 2015 by the authors; licensee MDPI, Basel, Switzerland. Source


Bai J.,Xiamen University | Gong W.,Xiamen University | Wang C.,Xiamen University | Gao Y.,Xiamen University | And 4 more authors.
Fish Physiology and Biochemistry | Year: 2015

In vertebrates, the aromatase coded by the cyp19a1a gene can catalyze the conversion from androgens to estrogens. Thus, the regulatory mechanisms of cyp19a1a gene expression are a critical research field in reproductive endocrinology. In this study, we use zebrafish as a model to study the dynamic methylation levels of the cyp19a1a gene core promoter during zebrafish ovarian folliculogenesis. The results show that there is an apparent fluctuation of the methylation levels of zebrafish cyp19a1a core promoter. Moreover, the methylation levels are inversely correlated with the expression levels of cyp19a1a transcripts when the ovarian follicles develop from PV into the MV stage. Also, the CpG dinucleotides which are close to the transcriptional starting site may have provided a significant blocking effect on inhibiting the transcriptional function of RNA polymerase II. Taken together, the results from the present study strongly suggest that DNA methylation was one of mechanisms that are involved in the regulation of cyp19a1a gene expression during folliculogenesis. This methylation mechanism modifying transcriptional process accompanied with zebrafish ovarian folliculogenesis might also shed new light on the regulation of cyp19a1a expression during the ovarian developmental stage in other vertebrates. © 2015 Springer Science+Business Media Dordrecht Source


Ao J.,State Oceanic Administration | Ao J.,Fujian Collaborative Innovation Center for Exploitation and Utilization of Marine Biological Resources | Ding Y.,State Oceanic Administration | Ding Y.,Fujian Collaborative Innovation Center for Exploitation and Utilization of Marine Biological Resources | And 7 more authors.
International Journal of Molecular Sciences | Year: 2015

The C-type lectin-like receptors (CTLRs) play important roles in innate immunity as one type of pattern recognition receptors. Here, we cloned and characterized a C-type lectin-like receptor (LycCTLR) from large yellow croaker Larimichthys crocea. The full-length cDNA of LycCTLR is 880 nucleotides long, encoding a protein of 215 amino acids. The deduced LycCTLR contains a C-terminal C-type lectin-like domain (CTLD), an N-terminal cytoplasmic tail, and a transmembrane region. The CTLD of LycCTLR possesses six highly conserved cysteine residues (C1–C6), a conserved WI/MGL motif, and two sugar binding motifs, EPD (Glu-Pro-Asp) and WYD (Trp-Tyr-Asp). Ca2+ binding site 1 and 2 were also found in the CTLD. The LycCTLR gene consists of five exons and four introns, showing the same genomic organization as tilapia (Oreochromis niloticus) and guppy (Poecilia retitculata) CTLRs. LycCTLR was constitutively expressed in various tissues tested, and its transcripts significantly increased in the head kidney and spleen after stimulation with inactivated trivalent bacterial vaccine. Recombinant LycCTLR (rLycCTLR) protein produced in Escherichia coli BL21 exhibited not only the hemagglutinating activity and a preference for galactose, but also the agglutinating activity against two food-borne pathogenic bacteria E. coli and Bacillus cereus in a Ca2+-dependent manner. These results indicate that LycCTLR is a potential galactose-binding C-type lectin that may play a role in the antibacterial immunity in fish. © 2015 by the authors; licensee MDPI, Basel, Switzerland. Source

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