Article
Biochemical Research Methods
Yasunori Sugiyama, Yuuki Uezato
Summary: Protein kinases play crucial roles in regulating biological processes and disease pathogenesis. Investigating the phosphorylation status of protein kinases is essential for understanding biological phenomena and disease mechanisms. Phos-tag, a tool for studying protein phosphorylation, along with techniques like Phos-tag SDS-PAGE, has been developed for analyzing protein phosphorylation.
JOURNAL OF PROTEOMICS
(2022)
Review
Biochemical Research Methods
Eiji Kinoshita, Emiko Kinoshita-Kikuta, Tohru Koike
Summary: Phos-tag is a functional molecule that selectively captures a phosphate monoester dianion in neutral aqueous solutions with an affinity more than 10,000 times greater than other anions present in living organisms. Developed techniques using Phos-tag have proven useful in phosphoproteomics, including separation and concentration of phosphopeptides and phosphoproteins, detection on various arrays, and electrophoresis for separation and detection of phosphoproteins.
JOURNAL OF PROTEOMICS
(2022)
Article
Biochemical Research Methods
Yuki Okawara, Hisashi Hirano, Ayuko Kimura, Natsumi Sato, Yuriko Hayashi, Makoto Osada, Takao Kawakami, Norihisa Ootake, Eiji Kinoshita, Kiyotaka Fujita
Summary: Phos-tag diagonal electrophoresis, a technique developed to precisely identify changes in phosphoprotein electrophoretic mobility, resolves the issue of determining mobility shifts by Mn2+ Phos-tag when using separate SDS-PAGE and Phos-tag SDS-PAGE. By providing SDS-PAGE and Phos-tag SDS-PAGE patterns on a single gel, this technique is useful for identifying phosphorylation states of various proteins.
JOURNAL OF PROTEOMICS
(2021)
Article
Nanoscience & Nanotechnology
Danqing Wang, Junfeng Huang, Haoran Zhang, Ting-Jia Gu, Lingjun Li
Summary: Protein phosphorylation is an important PTM involved in many cellular functions. This study developed a novel platform for phosphopeptide enrichment using a phosphorylated cotton-fiber-based Ti(IV)-IMAC material. The material showed high efficiency and selectivity in enriching phosphopeptides and demonstrated excellent sensitivity and specificity in complex biological samples.
ACS APPLIED MATERIALS & INTERFACES
(2023)
Article
Fisheries
Phattara-orn Havanapan, Suparat Taengchaiyaphum, Atchara Paemanee, Nuanwan Phungthanom, Sittiruk Roytrakul, Kallaya Sritunyalucksana, Chartchai Krittanai
Summary: The study showed that yellow head virus (YHV) replicates mainly in granular hemocytes in infected shrimp, with higher replication levels in GC. Comparative phosphoproteomic profiles revealed differences in the expression levels of phosphoproteins in YHV-infected GC, particularly with caspase-3 playing a crucial role in YHV replication and shrimp mortality.
FISH & SHELLFISH IMMUNOLOGY
(2021)
Review
Biochemical Research Methods
Hisashi Hirano, Jun Shirakawa
Summary: Phos-tag electrophoresis is an important method for analyzing the phosphorylation state of proteins. The recently developed Phos-tag diagonal electrophoresis has overcome the drawbacks of the traditional method and made it easier to determine protein phosphorylation state.
EXPERT REVIEW OF PROTEOMICS
(2022)
Review
Biochemical Research Methods
Emiko Kinoshita-Kikuta, Tohru Koike, Eiji Kinoshita
Summary: The Phos-tag technique is suitable for quantitative analysis of His- and Asp-phosphorylated proteins in a bacterial two-component system. This technique allows monitoring of auto-phosphorylation reactions of HK and RR, as well as phosphotransfer reactions from HK to RR. Additionally, the technique can be used for profiling potential HK inhibitors, providing a simple and convenient approach for drug discovery targeting the bacterial TCS.
JOURNAL OF PROTEOMICS
(2022)
Article
Biochemical Research Methods
Harunori Yoshikawa, Kohei Nishino, Hidetaka Kosako
Summary: In this study, novel ERK substrates were identified using a phosphoproteomic approach, and the phosphorylation of these substrates by active ERK was confirmed in both cellular and in vitro experiments. The study demonstrates the usefulness of Phos-tag SDS-PAGE for validating candidate substrates of protein kinases.
JOURNAL OF PROTEOMICS
(2022)
Article
Polymer Science
Sachio Yamamoto, Shoko Yano, Mitsuhiro Kinoshita, Shigeo Suzuki
Summary: An improved method for online preconcentration, derivatization, and separation of phosphorylated compounds was developed using a Phos-tag acrylamide gel in a polydimethylsiloxane/glass multichannel microfluidic chip. Phosphorylated peptides were specifically concentrated in the gel and derivatized with DTAF for sensitive analysis. This method enabled the sensitive analysis of phosphorylated peptides through a complex elution process.
Article
Plant Sciences
Keiji Nishioka, Yusuke Kato, Shin-ichiro Ozawa, Yuichiro Takahashi, Wataru Sakamoto
Summary: Protein phosphorylation plays a crucial role in regulating photosynthetic activities in organisms. The Phos-tag technology allows for a comprehensive assessment of protein phosphorylation in the thylakoid membrane of Arabidopsis, making it a useful biochemical tool for studying in vivo protein phosphorylation.
PHOTOSYNTHESIS RESEARCH
(2021)
Article
Biochemical Research Methods
Cho-Long Kim, Su -Bin Lim, Kyeongseob Kim, Han-Sol Jeong, Jung-Soon Mo
Summary: Phosphorylation is an essential regulatory mechanism in cells, influencing signal transduction and protein function. Dysregulation of phosphorylation can lead to diseases, making the study of protein phosphorylation crucial for understanding biological functions and drug discovery. Phos-tag technology can aid in the identification and quantitation of protein phosphorylation. This review provides insights into the application of Phos-tag techniques in studying phosphorylation of proteins in the Hippo pathway.
JOURNAL OF PROTEOMICS
(2022)
Article
Biochemical Research Methods
Yoko Ino, Mayuko Nishi, Yutaro Yamaoka, Kei Miyakawa, Sundararaj Stanleyraj Jeremiah, Makoto Osada, Yayoi Kimura, Akihide Ryo
Summary: This study investigated the functional phosphorylation of SARS-CoV-2 NP using Phos-tag technology. A unique phosphorylation site at Ser79 in NP was identified, and it was found that Pin1 positively regulated the production of viral particles by binding to phosphorylated NP at Ser79. These findings suggest that SARS-CoV-2 may have acquired a potent virus-host interaction mediated by viral protein phosphorylation during its evolution.
JOURNAL OF PROTEOMICS
(2022)
Article
Biochemistry & Molecular Biology
Gema Gonzalez-Rubio, Angela Sellers-Moya, Humberto Martin, Maria Molina
Summary: Slt2, a central MAPK in the yeast Cell Wall Integrity pathway, undergoes phosphorylation at Y192 and T190 sites, with Y192 phosphorylation being crucial for T190 phosphorylation. The activity of Slt2 is dependent on the phosphorylation status at these specific sites, highlighting the importance of precise detection of Slt2 phosphorylation status.
INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES
(2021)
Review
Biochemistry & Molecular Biology
Li-Rong Yu, Timothy D. Veenstra
Summary: Phosphorylation is a crucial post-translational modification for controlling physiological activities, and mass spectrometry has greatly facilitated the identification of phosphorylation sites in proteins, allowing for in-depth studies of protein phosphorylation.
CURRENT PROTEIN & PEPTIDE SCIENCE
(2021)
Article
Chemistry, Analytical
Lang Peng, Cunli Wang, Qiwen Lin, Yongxin Chang, Xiaoyu Zhang, Jing Wang, Zan Li, Zhiying Yang, Wenjing Sun, Wenqi Lu, Dongdong Wang, Guangyan Qing
Summary: Oxidation and protein phosphorylation are critical in regulating cellular activities. A new nanochannel device has been developed to detect both oxidation and protein phosphorylation using modified peptides. The device shows a sensitive response to reactive oxygen species and phosphorylated peptides, enabling the detection of cellular signaling pathways and kinase activity.
ANALYTICAL CHEMISTRY
(2023)
Article
Biochemical Research Methods
Hiroshi Kusamoto, Akio Shiba, Norinao Koretake, Haruto Fujioka, Yuhzo Hieda, Emiko Kinoshita-Kikuta, Eiji Kinoshita, Tohru Koike
JOURNAL OF CHROMATOGRAPHY B-ANALYTICAL TECHNOLOGIES IN THE BIOMEDICAL AND LIFE SCIENCES
(2016)
Article
Biochemical Research Methods
Eiji Kinoshita, Emiko Kinoshita-Kikuta, Yuji Kubota, Mutsuhiro Takekawa, Tohru Koike
Article
Chemistry, Analytical
Akio Shiba, Emiko Kinoshita-Kikuta, Eiji Kinoshita, Tohru Koike
Article
Biochemistry & Molecular Biology
Yuuki Uezato, Isamu Kameshita, Keiko Morisawa, Shuji Sakamoto, Eiji Kinoshita, Emiko Kinoshita-Kikuta, Tohru Koike, Yasunori Sugiyama
BIOCHIMICA ET BIOPHYSICA ACTA-PROTEINS AND PROTEOMICS
(2019)
Article
Biochemistry & Molecular Biology
Emiko Kinoshita-Kikuta, Eiji Kinoshita, Sayaka Ueda, Yoko Ino, Yayoi Kimura, Hisashi Hirano, Tohru Koike
BIOCHIMICA ET BIOPHYSICA ACTA-PROTEINS AND PROTEOMICS
(2019)
Article
Chemistry, Inorganic & Nuclear
Hiroshi Kusamoto, Akio Shiba, Masaya Tsunehiro, Haruto Fujioka, Emiko Kinoshita-Kikuta, Eiji Kinoshita, Tohru Koike
DALTON TRANSACTIONS
(2018)
Article
Biochemical Research Methods
Emiko Kinoshita-Kikuta, Hiroshi Kusamoto, Syogo Ono, Keisuke Akayama, Yoko Eguchi, Masayuki Igarashi, Toshihide Okajima, Ryutaro Utsumi, Eiji Kinoshita, Tohru Koike
Article
Multidisciplinary Sciences
Emiko Kinoshita-Kikuta, Ayane Tanikawa, Takuro Hosokawa, Aya Kiwado, Koko Moriya, Eiji Kinoshita, Tohru Koike, Toshihiko Utsumi
Article
Biochemical Research Methods
Masaya Tsunehiro, Kenji Sasaki, Emiko Kinoshita-Kikuta, Eiji Kinoshita, Tohru Koike
JOURNAL OF CHROMATOGRAPHY B-ANALYTICAL TECHNOLOGIES IN THE BIOMEDICAL AND LIFE SCIENCES
(2020)
Article
Biochemical Research Methods
Emiko Kinoshita-Kikuta, Shino Maruta, Yoko Eguchi, Masayuki Igarashi, Toshihide Okajima, Ryutaro Utsumi, Eiji Kinoshita, Tohru Koike
ANALYTICAL BIOCHEMISTRY
(2020)
Article
Multidisciplinary Sciences
Emiko Kinoshita-Kikuta, Toshihiko Utsumi, Aya Miyazaki, Chiharu Tokumoto, Kyosuke Doi, Haruna Harada, Eiji Kinoshita, Tohru Koike
SCIENTIFIC REPORTS
(2020)
Article
Biochemical Research Methods
Emiko Kinoshita-Kikuta, Momoka Yoshimoto, Marina Yano, Eiji Kinoshita, Tohru Koike
Summary: This article discusses an assay of tyrosine protein kinase ABL activity using an Escherichia coli protein expression system. The study involves co-expressing human tyrosine kinase ABL and its substrate in E. coli and detecting tyrosine phosphorylation using Phos-tag SDS-PAGE. Results show different kinase activity levels in mutant forms of ABL compared to the wild-type, indicating a correlation between phosphorylation states and enzymatic activity.
Article
Biochemistry & Molecular Biology
Emiko Kinoshita-Kikuta, Eiji Kinoshita, Misaki Suga, Mana Higashida, Yuka Yamane, Tomoka Nakamura, Tohru Koike
Summary: The production of heterologous proteins is important for biologists, with various cell-based and cell-free systems available. This study prepared human Src family kinases using six different expression systems and analyzed their phosphorylation status and activities. Eukaryotic systems showed multiple phosphorylated forms, while prokaryotic systems expressed active forms through autophosphorylation.
Article
Biochemical Research Methods
Eiji Kinoshita, Emiko Kinoshita-Kikuta, Kiyonobu Karata, Toshiki Kawano, Atsuhiro Nishiyama, Morihisa Yamato, Tohru Koike
Article
Biochemical Research Methods
Elena Tianfei Yuan, Yoko Ino, Maho Kawaguchi, Yayoi Kimura, Hisashi Hirano, Emiko Kinoshita-Kikuta, Eiji Kinoshita, Tohru Koike