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Journal of Biochemistry Advance Access published online on May 18, 2009
Journal of Biochemistry, doi:10.1093/jb/mvp075
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© The authors 2009. Published by Oxford University Press on behalf of the Japanese Biochemical Society. All rights reserved.

NMR structure of the heterodimer of Bem1 and Cdc24 PB1 domains from Saccharomyces cerevisiae

Kenji Ogura1,*, Tsubasa Tandai1,*, Sosuke Yoshinaga1,*, Yoshihiro Kobashigawa1, Hiroyuki Kumeta1, Takashi Ito2, Hideki Sumimoto3 and Fuyuhiko Inagaki1

1Department of Structural Biology, Graduate School of Pharmaceutical Sciences, Hokkaido University, Kita 12 Nishi 6, Kita-ku, Sapporo 060-0812, Japan.
2Department of Computational Biology, Graduate School of Frontier Sciences, University of Tokyo, 5-1-5 Kashiwanoha, Kashiwa 277-8561, Japan.
3Medical Institute of Bioregulation, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka 812-8582, Japan.

To whom correspondence should be addressed: Prof. Fuyuhiko Inagaki Department of Structural Biology, Graduate School of Pharmaceutical Sciences, Hokkaido University Kita 12 Nishi 6, Kita, Sapporo 060-0812, Japan. Tel: +81-11-706-9011 FAX: +81-11-706-9012 Email: finagaki{at}pharm.hokudai.ac.jp

Received April 2, 2009; Accepted April 30, 2009


  Abstract

Bem1 and Cdc24 of the budding yeast Saccharomyces cerevisiae interact with each other through PB1-PB1 heterodimer formation to regulate the establishment of cell polarity. Here we present the tertiary structure of the heterodimer of Bem1 and Cdc24 PB1 domains determined by NMR spectroscopy. To avoid ambiguity in the NMR spectral analysis, we first prepared a mutant of the Cdc24 PB1 domain that had truncated loops. The mutant provided well dispersed spectra without spectral overlapping, thus allowing unambiguous spectral assignments for structure determination. We confirmed that the loop deletion-mutant was quite similar to the wild type in both three-dimensional structure and binding affinity. The NMR structure of the heterodimer of the deletion-mutant of Cdc24 PB1 and Bem1 PB1 was determined using a variety of isotope labeled samples including perdeuteration.?The interface between the Bem1/Cdc24 PB1 heterodimer was analyzed at atomic resolution. Through a comparison with the tertiary structures of other PB1-PB1 heterodimers, we found that conserved electrostatic properties on the molecular surface were commonly used for PB1-PB1 interaction, but hydrophobic interactions were important for cognate interaction in Bem1/Cdc24 PB1 heterodimer formation.

Key Words: Bem1, Cdc24, PB1 domain, heterodimer, NMR, perdeutration, residual dipolar coupling, solution structure

* These authors contributed equally to this work.


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