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Observation of the Membrane Binding Activity and Domain Structure of gpV, Which Comprises the Tail Spike of Bacteriophage P2

Yasuhiro Kageyama‡, Masanori Murayama‡, Takashi Onodera‡, Seiko Yamada‡, Harumi Fukada§, Motonori Kudou

, Kouhei Tsumoto

, Yoshiharu Toyama‡, Syunsaku Kado‡, Kenji Kubota‡ and Shigeki Takeda*‡

^‡ Department of Chemical Biology, Graduate School of Engineering, Gunma University, 1-5-1 Tenjin-cho, Kiryu, Gunma 376-8515, Japan

^§ Graduate School of Agriculture and Biological Sciences, Osaka Prefecture University, Sakai, Osaka 599-8531, Japan

Department of Medical Genome Sciences, Graduate School of Frontier Sciences, The University of Tokyo, 301 FBS-Building, 5-1-5 Kashiwanoha, Kashiwa 277-8562, Japan

Biochemistry, 2009, 48 (42), pp 10129–10135

DOI: 10.1021/bi900928n

Publication Date (Web): September 25, 2009

*To whom correspondence should be addressed: Department of Chemical Biology, Graduate School of Engineering, Gunma University, 1-5-1 Tenjin-cho, Kiryu, Gunma 376-8515, Japan. Telephone or fax: +81-277-30-1434. E-mail: stakeda@chem-bio.gunma-u.ac.jp.

Abstract

The P2 phage virion has tail spike proteins beneath the baseplate and uses them to adsorb to the outer membrane of Escherichia coli during the infection process. Previous immunoelectron microscopic studies suggested that the tail spikes are composed of the gene V product (gpV); however, experimental evidence of its membrane binding activity has yet to be reported. In this study, we purified and characterized recombinant full-length gpV and its C-terminal domain. Limited chymotrypsin proteolysis of gpV produced a C-terminal domain composed of Ser86−Leu211. Our experiments demonstrated that the N- and C-terminal domains have very different melting temperatures: 50 and 74 °C, respectively. We also found that gpV binds the E. coli membrane via its C-terminal domain. We conclude that the C-terminal domain of gpV is a stable trimer and serves as the receptor-binding domain for the second step in the phage adsorption process.

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History

Published In Issue October 27, 2009
Article ASAPOctober 02, 2009
Just Accepted ManuscriptSeptember 25, 2009
Received: June 02, 2009
Revised: September 23, 2009

Article

Observation of the Membrane Binding Activity and Domain Structure of gpV, Which Comprises the Tail Spike of Bacteriophage P2