The Baseplate Wedges of Bacteriophage T4 Spontaneously Assemble into Hubless Baseplate-Like Structure In Vitro

Moh Lan Yap, Kazuhiro Mio, Petr Leiman, Shuji Kanamaru, Fumio Arisaka

Research output: Contribution to journalArticle

24 Citations (Scopus)

Abstract

The baseplate of phage T4 is an important model system in viral supramolecular assembly. The baseplate consists of six wedges surrounding the central hub. We report the first successful attempt at complete wedge assembly using an in vitro approach based on recombinant proteins. The cells expressing the individual wedge proteins were mixed in a combinatorial manner and then lysed. Using this approach, we could both reliably isolate the complete wedge along with a series of intermediate complexes as well as determine the exact sequence of assembly. The individual proteins and intermediate complexes at each step of the wedge assembly were successfully purified and characterized by sedimentation velocity and electron microscopy. Although our results mostly confirmed the hypothesized sequential wedge assembly pathway as established using phage mutants, interestingly, we also detected some protein interactions not following the specified order. It was found that association of gene product 53 to the immediate precursor complex induces spontaneous association of the wedges to form a six-fold star-shaped baseplate-like structure in the absence of the hub. The formation of the baseplate-like structure was facilitated by the addition of gene product 25. The complete wedge in the star-shaped supramolecular complex has a structure similar to the baseplate in the expanded "star" conformation found after infection. Based on the results of the present and previous studies, we assume that the strict order of wedge assembly is due to the induced conformational change caused by every new binding event. The significance of a 40-S star-shaped baseplate structure, which was previously reported and was also found in this study, is discussed in the light of a new paradigm for T4 baseplate assembly involving the star-shaped wedge ring and the central hub. Importantly, the methods described in this article suggest a novel methodology for future structural characterization of supramolecular protein assemblies.

Original languageEnglish (US)
Pages (from-to)349-360
Number of pages12
JournalJournal of Molecular Biology
Volume395
Issue number2
DOIs
StatePublished - Jan 15 2010
Externally publishedYes

Fingerprint

Bacteriophage T4
Proteins
Virus Assembly
Recombinant Proteins
Bacteriophages
Genes
Electron Microscopy
In Vitro Techniques
Infection

Keywords

  • analytical ultracentrifugation
  • assembly
  • bacteriophage T4
  • baseplate
  • electron microscopy

ASJC Scopus subject areas

  • Molecular Biology

Cite this

The Baseplate Wedges of Bacteriophage T4 Spontaneously Assemble into Hubless Baseplate-Like Structure In Vitro. / Yap, Moh Lan; Mio, Kazuhiro; Leiman, Petr; Kanamaru, Shuji; Arisaka, Fumio.

In: Journal of Molecular Biology, Vol. 395, No. 2, 15.01.2010, p. 349-360.

Research output: Contribution to journalArticle

Yap, Moh Lan ; Mio, Kazuhiro ; Leiman, Petr ; Kanamaru, Shuji ; Arisaka, Fumio. / The Baseplate Wedges of Bacteriophage T4 Spontaneously Assemble into Hubless Baseplate-Like Structure In Vitro. In: Journal of Molecular Biology. 2010 ; Vol. 395, No. 2. pp. 349-360.
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