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Kota Mayanagi Last modified date:2018.07.18



Other Organization


Homepage
http://www.bioreg.kyushu-u.ac.jp/vsb/index.html
Division of Structural Biology, Institute of Bioregulation, Kyushu University .
Academic Degree
PhD
Field of Specialization
Structural Biology
Outline Activities
Three Dimensional Electron Microscopy of Macromolecules
Research
Research Interests
  • Electron microscopic single particle analysis of DNA-protein complexes
    keyword : Electron microscopy, Single particle analysis, Macromolecule, Image analysis, Structral Biology
    1991.04.
Academic Activities
Papers
1. Mayanagi K, Ishino S Takafuji M, Mitsuoka K, Shirai T, Kiyonari S, Nishida H, Kohda D, Morikawa K, Ishino Y., Switching mechanism of DNA replication fork complex revealed by single particle analysis, European Microscopy Congress , 3, 57-58, Vol3, 57-58, 2016.12.
2. Aramaki S, Mayanagi K, Jin M, Aoyama K, Yasunaga T, Filopodia formation by crosslinking of F-actin with fascin in two different binding manners, Cytoskeleton, 10.1002/cm.21309, 73(7):365-74, 2016.06.
3. Takekawa N, Terahara N, Kato T, Gohara M, Mayanagi K, Hijikata A, Onoue Y, Kojima S, Shirai T, Namba K, Homma M, The tetrameric MotA complex as the core of the flagellar motor stator from hyperthermophilic bacterium, Scientific Reports, 10.1038/srep31526, 6:31526, 2016.08.
4. Nakae S, Hijikata A, Tsuji T, Yonezawa K, Kouyama KI, Mayanagi K, Ishino S, Ishino Y, Shirai T, Structure of the EndoMS-DNA Complex as Mismatch Restriction Endonuclease, Structure, 24, 1960-1971, 24, 1960-1971, 2016.07, Archaeal NucS nuclease was thought to degrade the single-stranded region of branched DNA, which contains flapped and splayed DNA. However, recent findings indicated that EndoMS, the orthologous enzyme of NucS, specifically cleaves double-stranded DNA (dsDNA) containing mismatched bases. In this study, we determined the structure of the EndoMS-DNA complex. The complex structure of the EndoMS dimer with dsDNA unexpectedly revealed that the mismatched bases were flipped out into binding sites, and the overall architecture most resembled that of restriction enzymes. The structure of the apo form was similar to the reported structure of Pyrococcus abyssi NucS, indicating that movement of the C-terminal domain from the resting state was required for activity. In addition, a model of the EndoMS-PCNA-DNA complex was preliminarily verified with electron microscopy. The structures strongly support the idea that EndoMS acts in a mismatch repair pathway..
5. Mayanagi K, Kiyonari S, Nishida H, Saito M, Kohda D, Ishino Y, Shirai T, Morikawa K, , Architecture of the DNA polymerase B-proliferating cell nuclear antigen (PCNA)-DNA ternary complex. , PNAS, 108, 5, 1845-1849, (Corresponding Author), 2011.02.
6. Hirokazu. Nishida, Kouta. Mayanagi, Shinichi. Kiyonari, Yuichi. Sato, Takuji. Oyama, Yoshizumi. Ishino, Kosuke. Morikawa, Structural determinant for switching between the polymerase and exonuclease modes in the PCNA-replicative DNA polymerase complex, PNAS, 106, 20693-20698, 2009.11.
7. Kouta Mayanagi, Shinichi Kiyonari, Mihoko Saito, Tsuyoshi Shirai, Yoshizumi Ishino, Kosuke Morikawa, Mechanism of replication machinery assembly as revealed by the DNA ligase-PCNA-DNA complex architecture, PNAS, 106, 4647-4652, (Corresponding Author), 2009.03.
8. Yasuto Murayama, Yumiko Kurokawa, Kouta Mayanagi, and Hiroshi Iwasaki, Formation and branch migration of Holliday junctions mediated by eukaryotic recombinases, Nature, 451, 7181, 1018-1021, 2008.02.
9. Yoshie Fujiwara, Kouta Mayanagi, and Kosuke Morikawa, Functional significance of octameric RuvA for a branch migration complex from Thermus thermophilus., BBRC, 366, 2, 426-431, 2007.12.
10. Kouta Mayanagi, Yoshie Fujiwara, Tomoko Miyata, and Kosuke Morikawa, Electron microscopic single particle analysis of a tetrameric RuvA/RuvB/Holliday junction DNA complex, BBRC, 365, 273-278, (Corresponding Author), 2007.11.
11. Tomoko Miyata, Hirofumi Suzuki, Takuji Oyama, Kouta Mayanagi, Yoshizumi Ishino, and Kosuke Morikawa, Open clamp structure in the clamp-loading complex visualized by electron microscopic image analysis. , PNAS, 10.1073/pnas.0506447102, 102, 39, 13795-13800, 2005.09.
12. Tomoko Miyata, Takuji Oyama, Kouta Mayanagi, Sonoko Ishino, Yoshizumi Ishino, and Kosuke Morikawa, The Clamp-loading Complex for Processive DNA Replication, Nature Struct. & Molec. Biol. , 10.1038/nsmb788, 11, 7, 632-636, 2004.06.
13. Kyoko Matoba, Kouta Mayanagi, Syo Nakasu, Akihiko Kikuchi, and Kosuke Morikawa, Three-dimensional electron microscopy of the reverse gyrase from Sulfolobus tokodaii, BBRC, 297, 4, 749-755, 2002.10.
14. Naoko Kajimura, Matsuyo Yamazaki, Kosuke Morikawa, Akio Yamazaki, and Kouta Mayanagi, Three-dimensional structure of non-activated cGMP phosphodiesterase 6 and comparison of its image with those of activated forms, J. Struct. Biol., 139, 1, 27-38, (Corresponding Author), 2002.07.
15. Mayanagi, K., Miyata, T., Oyama, T., Ishino, Y., and Morikawa, K, Three-dimensional electron microscopy of the clamp loader small subunit from Pyrococcus furiosus, J. Struct. Biol., 134, 1, 35-45, (Corresponding Author), 2001.04.
16. Miyata, T., Yamada, K., Iwasaki, H., Shinagawa, H., Morikawa, K., and Mayanagi. K, Two different oligomeric states of the RuvB branch migration motor protein as revealed by electron microscopy, J. Struct. Biol., 131, 2, 83-89, (Corresponding Author), 2000.08.
17. Mayanagi, K., Ishikawa, T., Toyoshima, C., Inoue, Y., and Nakazato, K, Three-dimensional electron microscopy of the photosystem II core complex, J. Struct. Biol. , 123, 3, 211-224, 1998.11.
Presentations
1. Kouta Mayanagi, Single particle analysis of DNA replication fork complex, 10th International 3R (DNA, Replication, Recombination, and Repair) Symposium, 2016.11,  We have been studying the mechanism of synthesis and maturation of Okazaki fragments, in which three replication factors, i.e. DNA polymerase, Flap endonuclease (FEN), and DNA ligase, are playing essential roles. As all of these three enzymes are known to interact with PCNA, a switching mechanism between these factors, bound simultaneously to a PCNA ring has been proposed, called PCNA tool belt model, which is considered to increase the efficiency of these sequential reactions. Recent biochemical studies, however, suggest sequential binding and releasing of these factors. Little is known regarding the precise switching mechanism, due to the lack of the structural knowledge of these complexes.
We investigated the structures of the core components of the replisome, such as DNA polymerase-PCNA-DNA, and DNA ligase-PCNA-DNA, by single particle analysis. Besides the known interaction through a PIP-box motif, we could find a novel contact between polymerase and PCNA, as well as between ligase and PCNA. Mutant analysis showed that these contacts are involved in the regulation of the replication factors, such as the switching between the polymerizing and editing modes of the polymerase. Our results, showing that both factors are bound to two subunits of the PCNA trimer, were inconsistent with the standard tool belt model. The third PCNA subunit, however, was still free in both complexes, thus we analyzed the complex structures with two replication factors. We could successfully visualize the 3D structure of FEN-DNA ligase-PCNA-DNA complex, and the handing over of the ds DNA from FEN to DNA ligase..
2. Mayanagi K, Ishino S Takafuji M, Mitsuoka K, Shirai T, Kiyonari S, Nishida H, Kohda D, Morikawa K, Ishino Y., Switching mechanism of the replisome as revealed by single particle analysis, 第 89 回日本生化学会大会, 2016.09, DNA replication in archaea and eukaryotes is executed by family B DNA polymerases, which exhibitfull activity when complexed with the DNA clamp, proliferating cell nuclear antigen (PCNA). PCNAhas a trimeric ring structure that encircles the DNA, and increases the processivity of the bound DNApolymerase by tethering it to the DNA. It is known now, that PCNA also interacts with multiple partnersto control DNA replication, DNA repair, and cell cycle progression, and works not only as the platform,but also as the conductor for the recruitment and release of these factors. However, the moleculararchitectures as well as the mechanism of the regulation of these replication factors are not known indetail.We have investigated the three-dimensional structure of the core components of the replisome,such as DNA polymerase-PCNA-DNA, and DNA ligase-PCNA-DNA ternary complexes, by singleparticle analysis (2- 3). Besides the authentic interaction through a PCNA-interacting protein box (PIPbox),we could find a novel contact between both polymerase-PCNA and ligase-PCNA. Mutant analysisshowed that these contacts are involved in the regulation of the replication factors, such as the switchingbetween the polymerizing and editing modes of the polymerase. Our results, showing that both factorsbound to two subunits of the PCNA trimer ring, were inconsistent with the standard tool belt model. Thethird PCNA subunit, however, was still free in both complexes, thus we analyzed the complex structureswith two replication factors bound to one PCNA ring, in order to investigate the switching mechanismbetween them in more detail..
3. Mayanagi K, Ishino S Takafuji M, Mitsuoka K, Shirai T, Kiyonari S, Nishida H, Kohda D, Morikawa K, Ishino Y., Switching mechanism of DNA replication fork complex revealed by single particle analysis, The 16th European Microscopy Congress, 2016.08.
4. Kouta Mayanagi, Molecular switching of DNA replication fork complex revealed by single particle analysis, The 1st APPA-PSSJ joint workshop, 2016.06, Okazaki fragments synthesis and maturation, during lagging strand DNA replication, is accomplished by mainly three replication factors, i.e. DNA polymerase, Flap endonuclease, and DNA ligase. All of these three enzymes are known to interact with the DNA clamp, proliferating cell nuclear antigen (PCNA), through a PCNA-interacting protein box (PIP-box) motif. PCNA forms a trimeric ring that encircles the DNA, and increases the processivity of the bound DNA polymerase by tethering it to the DNA. As up to three replication factors could be bound to a PCNA trimer, a switching mechanism between these replication factors, bound simultaneously to a PCNA ring has been proposed, called PCNA tool belt model, which is considered to increase the efficiency of the sequential reactions. However, the precise mechanism is not known in detail, due to the lack of structural data of such complexes. In order to investigate the mechanism of the regulation and switching of these replication factors bound to PCNA, we have studied the molecular architectures of the core components of the replisome, such as DNA polymerase-PCNA-DNA, and DNA ligase-PCNA-DNA complexes, by single particle analysis..
5. Kouta Mayanagi, Electron microscopic analysis of molecular architecture and switching mechanism of DNA replication fork complex, ICSG2013-SLS, 2013.07.
6. Kouta Mayanagi, Shinichi Kiyonari, Hirokazu Nishida, Sonoko Iahino, Mihoko Saito, Daisuke Kohda, Yoshizumi Ishino, Tsuyoshi Shirai, Kosuke Morikawa, Molecular architecture and switching mechanism of DNA replication fork complex, Gordon Research Conferences: Three Dimensional Electron Microscopy, 2013.06.
Educational
Other Educational Activities
  • 2012.09.
  • 2014.08.
  • 2013.03.
  • 2012.03.
  • 2013.03.
  • 2014.03.
  • 2015.03.
  • 2018.03.
  • 2017.09.
  • 2017.03.
  • 2011.03.
  • 2011.03.