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Identification and Functional Analysis of Tartrate-Resistant Acid Phosphatase Type 5b (TRAP5b) in Oreochromis niloticus
Tartrate-resistant acid phosphatase type 5 (TRAP5) is an enzyme that is highly expressed in activated macrophages and osteoclasts and plays important biological functions in mammalian immune defense systems. In the study, we investigated the functions of tartrate-resistant acid phosphatase type 5b f...
Autores principales: | , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
MDPI
2023
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10138680/ https://www.ncbi.nlm.nih.gov/pubmed/37108342 http://dx.doi.org/10.3390/ijms24087179 |
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author | Lei, Yang Fu, Shengli Yang, Yanjian Chen, Jianlin Li, Bingxi Guo, Zheng Ye, Jianmin |
author_facet | Lei, Yang Fu, Shengli Yang, Yanjian Chen, Jianlin Li, Bingxi Guo, Zheng Ye, Jianmin |
author_sort | Lei, Yang |
collection | PubMed |
description | Tartrate-resistant acid phosphatase type 5 (TRAP5) is an enzyme that is highly expressed in activated macrophages and osteoclasts and plays important biological functions in mammalian immune defense systems. In the study, we investigated the functions of tartrate-resistant acid phosphatase type 5b from Oreochromis niloticus (OnTRAP5b). The OnTRAP5b gene has an open reading frame of 975 bp, which encodes a mature peptide consisting of 302 amino acids with a molecular weight of 33.448 kDa. The OnTRAP5b protein contains a metallophosphatase domain with metal binding and active sites. Phylogenetic analysis revealed that OnTRAP5b is clustered with TRAP5b of teleost fish and shares a high amino acid sequence similarity with other TRAP5b in teleost fish (61.73–98.15%). Tissues expression analysis showed that OnTRAP5b was most abundant in the liver and was also widely expressed in other tissues. Upon challenge with Streptococcus agalactiae and Aeromonas hydrophila in vivo and in vitro, the expression of OnTRAP5b was significantly up-regulated. Additionally, the purified recombinant OnTRAP5b ((r)OnTRAP5) protein exhibited optimal phosphatase activity at pH 5.0 and an ideal temperature of 50 °C. The V(max), K(m,) and k(cat) of purified (r)OnTRAP5b were found to be 0.484 μmol × min(−1) × mg(−1), 2.112 mM, and 0.27 s(−1) with respect to pNPP as a substrate, respectively. Its phosphatase activity was differentially affected by metal ions (K(+), Na(+), Mg(2+), Ca(2+), Mn(2+), Cu(2+), Zn(2+), and Fe(3+)) and inhibitors (sodium tartrate, sodium fluoride, and EDTA). Furthermore, (r)OnTRAP5b was found to promote the expression of inflammatory-related genes in head kidney macrophages and induce reactive oxygen expression and phagocytosis. Moreover, OnTRAP5b overexpression and knockdown had a significant effect on bacterial proliferation in vivo. When taken together, our findings suggest that OnTRAP5b plays a significant role in the immune response against bacterial infection in Nile tilapia. |
format | Online Article Text |
id | pubmed-10138680 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2023 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
spelling | pubmed-101386802023-04-28 Identification and Functional Analysis of Tartrate-Resistant Acid Phosphatase Type 5b (TRAP5b) in Oreochromis niloticus Lei, Yang Fu, Shengli Yang, Yanjian Chen, Jianlin Li, Bingxi Guo, Zheng Ye, Jianmin Int J Mol Sci Article Tartrate-resistant acid phosphatase type 5 (TRAP5) is an enzyme that is highly expressed in activated macrophages and osteoclasts and plays important biological functions in mammalian immune defense systems. In the study, we investigated the functions of tartrate-resistant acid phosphatase type 5b from Oreochromis niloticus (OnTRAP5b). The OnTRAP5b gene has an open reading frame of 975 bp, which encodes a mature peptide consisting of 302 amino acids with a molecular weight of 33.448 kDa. The OnTRAP5b protein contains a metallophosphatase domain with metal binding and active sites. Phylogenetic analysis revealed that OnTRAP5b is clustered with TRAP5b of teleost fish and shares a high amino acid sequence similarity with other TRAP5b in teleost fish (61.73–98.15%). Tissues expression analysis showed that OnTRAP5b was most abundant in the liver and was also widely expressed in other tissues. Upon challenge with Streptococcus agalactiae and Aeromonas hydrophila in vivo and in vitro, the expression of OnTRAP5b was significantly up-regulated. Additionally, the purified recombinant OnTRAP5b ((r)OnTRAP5) protein exhibited optimal phosphatase activity at pH 5.0 and an ideal temperature of 50 °C. The V(max), K(m,) and k(cat) of purified (r)OnTRAP5b were found to be 0.484 μmol × min(−1) × mg(−1), 2.112 mM, and 0.27 s(−1) with respect to pNPP as a substrate, respectively. Its phosphatase activity was differentially affected by metal ions (K(+), Na(+), Mg(2+), Ca(2+), Mn(2+), Cu(2+), Zn(2+), and Fe(3+)) and inhibitors (sodium tartrate, sodium fluoride, and EDTA). Furthermore, (r)OnTRAP5b was found to promote the expression of inflammatory-related genes in head kidney macrophages and induce reactive oxygen expression and phagocytosis. Moreover, OnTRAP5b overexpression and knockdown had a significant effect on bacterial proliferation in vivo. When taken together, our findings suggest that OnTRAP5b plays a significant role in the immune response against bacterial infection in Nile tilapia. MDPI 2023-04-13 /pmc/articles/PMC10138680/ /pubmed/37108342 http://dx.doi.org/10.3390/ijms24087179 Text en © 2023 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/). |
spellingShingle | Article Lei, Yang Fu, Shengli Yang, Yanjian Chen, Jianlin Li, Bingxi Guo, Zheng Ye, Jianmin Identification and Functional Analysis of Tartrate-Resistant Acid Phosphatase Type 5b (TRAP5b) in Oreochromis niloticus |
title | Identification and Functional Analysis of Tartrate-Resistant Acid Phosphatase Type 5b (TRAP5b) in Oreochromis niloticus |
title_full | Identification and Functional Analysis of Tartrate-Resistant Acid Phosphatase Type 5b (TRAP5b) in Oreochromis niloticus |
title_fullStr | Identification and Functional Analysis of Tartrate-Resistant Acid Phosphatase Type 5b (TRAP5b) in Oreochromis niloticus |
title_full_unstemmed | Identification and Functional Analysis of Tartrate-Resistant Acid Phosphatase Type 5b (TRAP5b) in Oreochromis niloticus |
title_short | Identification and Functional Analysis of Tartrate-Resistant Acid Phosphatase Type 5b (TRAP5b) in Oreochromis niloticus |
title_sort | identification and functional analysis of tartrate-resistant acid phosphatase type 5b (trap5b) in oreochromis niloticus |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10138680/ https://www.ncbi.nlm.nih.gov/pubmed/37108342 http://dx.doi.org/10.3390/ijms24087179 |
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