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Establishment of Novel High-Standard Chemiluminescent Assay for NTPase in Two Protozoans and Its High-Throughput Screening

Toxoplasma gondii is a major protozoan parasite and infects human and many other warm-blooded animals. The infection leads to Toxoplasmosis, a serious issue in AIDS patients, organ transplant recipients and pregnant women. Neospora caninum, another type of protozoa, is closely related to Toxoplasma...

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Autores principales: Harada, Masamitsu, Nagai, Jun, Kurata, Riho, Shimizu, Kenji, Cui, Xiaofeng, Isagawa, Takayuki, Semba, Hiroaki, Ishihara, Jun, Yoshida, Yasuhiro, Takeda, Norihiko, Maemura, Koji, Yonezawa, Tomo
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2020
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Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7142789/
https://www.ncbi.nlm.nih.gov/pubmed/32183121
http://dx.doi.org/10.3390/md18030161
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author Harada, Masamitsu
Nagai, Jun
Kurata, Riho
Shimizu, Kenji
Cui, Xiaofeng
Isagawa, Takayuki
Semba, Hiroaki
Ishihara, Jun
Yoshida, Yasuhiro
Takeda, Norihiko
Maemura, Koji
Yonezawa, Tomo
author_facet Harada, Masamitsu
Nagai, Jun
Kurata, Riho
Shimizu, Kenji
Cui, Xiaofeng
Isagawa, Takayuki
Semba, Hiroaki
Ishihara, Jun
Yoshida, Yasuhiro
Takeda, Norihiko
Maemura, Koji
Yonezawa, Tomo
author_sort Harada, Masamitsu
collection PubMed
description Toxoplasma gondii is a major protozoan parasite and infects human and many other warm-blooded animals. The infection leads to Toxoplasmosis, a serious issue in AIDS patients, organ transplant recipients and pregnant women. Neospora caninum, another type of protozoa, is closely related to Toxoplasma gondii. Infections of the protozoa in animals also causes serious diseases such as Encephalomyelitis and Myositis-Polyradiculitis in dogs or abortion in cows. Both Toxoplasma gondii and Neospora caninum have similar nucleoside triphosphate hydrolases (NTPase), NcNTPase and TgNTPase-I in Neospora caninum and Toxoplasma gondii, respectively. These possibly play important roles in propagation and survival. Thus, we targeted the enzymes for drug discovery and tried to establish a novel high-standard assay by a combination of original biochemical enzyme assay and fluorescent assay to determine ADP content. We then validated whether or not it can be applied to high-throughput screening (HTS). Then, it fulfilled criterion to carry out HTS in both of the enzymes. In order to identify small molecules having inhibitory effects on the protozoan enzyme, we also performed HTS using two synthetic compound libraries and an extract library derived from marine bacteria and then, identified 19 compounds and 6 extracts. Nagasaki University collected many extracts from over 18,000 marine bacteria found in local Omura bay, and continues to compile an extensive collection of synthetic compounds from numerous drug libraries established by Japanese chemists.
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spelling pubmed-71427892020-04-14 Establishment of Novel High-Standard Chemiluminescent Assay for NTPase in Two Protozoans and Its High-Throughput Screening Harada, Masamitsu Nagai, Jun Kurata, Riho Shimizu, Kenji Cui, Xiaofeng Isagawa, Takayuki Semba, Hiroaki Ishihara, Jun Yoshida, Yasuhiro Takeda, Norihiko Maemura, Koji Yonezawa, Tomo Mar Drugs Article Toxoplasma gondii is a major protozoan parasite and infects human and many other warm-blooded animals. The infection leads to Toxoplasmosis, a serious issue in AIDS patients, organ transplant recipients and pregnant women. Neospora caninum, another type of protozoa, is closely related to Toxoplasma gondii. Infections of the protozoa in animals also causes serious diseases such as Encephalomyelitis and Myositis-Polyradiculitis in dogs or abortion in cows. Both Toxoplasma gondii and Neospora caninum have similar nucleoside triphosphate hydrolases (NTPase), NcNTPase and TgNTPase-I in Neospora caninum and Toxoplasma gondii, respectively. These possibly play important roles in propagation and survival. Thus, we targeted the enzymes for drug discovery and tried to establish a novel high-standard assay by a combination of original biochemical enzyme assay and fluorescent assay to determine ADP content. We then validated whether or not it can be applied to high-throughput screening (HTS). Then, it fulfilled criterion to carry out HTS in both of the enzymes. In order to identify small molecules having inhibitory effects on the protozoan enzyme, we also performed HTS using two synthetic compound libraries and an extract library derived from marine bacteria and then, identified 19 compounds and 6 extracts. Nagasaki University collected many extracts from over 18,000 marine bacteria found in local Omura bay, and continues to compile an extensive collection of synthetic compounds from numerous drug libraries established by Japanese chemists. MDPI 2020-03-13 /pmc/articles/PMC7142789/ /pubmed/32183121 http://dx.doi.org/10.3390/md18030161 Text en © 2020 by the authors. 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 (http://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Harada, Masamitsu
Nagai, Jun
Kurata, Riho
Shimizu, Kenji
Cui, Xiaofeng
Isagawa, Takayuki
Semba, Hiroaki
Ishihara, Jun
Yoshida, Yasuhiro
Takeda, Norihiko
Maemura, Koji
Yonezawa, Tomo
Establishment of Novel High-Standard Chemiluminescent Assay for NTPase in Two Protozoans and Its High-Throughput Screening
title Establishment of Novel High-Standard Chemiluminescent Assay for NTPase in Two Protozoans and Its High-Throughput Screening
title_full Establishment of Novel High-Standard Chemiluminescent Assay for NTPase in Two Protozoans and Its High-Throughput Screening
title_fullStr Establishment of Novel High-Standard Chemiluminescent Assay for NTPase in Two Protozoans and Its High-Throughput Screening
title_full_unstemmed Establishment of Novel High-Standard Chemiluminescent Assay for NTPase in Two Protozoans and Its High-Throughput Screening
title_short Establishment of Novel High-Standard Chemiluminescent Assay for NTPase in Two Protozoans and Its High-Throughput Screening
title_sort establishment of novel high-standard chemiluminescent assay for ntpase in two protozoans and its high-throughput screening
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7142789/
https://www.ncbi.nlm.nih.gov/pubmed/32183121
http://dx.doi.org/10.3390/md18030161
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