Altering the speed of a DNA packaging motor from bacteriophage T4

Siying Lin, Tanfis I. Alam, Vishal I. Kottadiel, Carl J. Vangessel, Wei Chun Tang, Yann R. Chemla, Venigalla B. Rao

Research output: Contribution to journalArticle

Abstract

The speed at which a molecular motor operates is critically important for the survival of a virus or an organism but very little is known about the underlying mechanisms. Tailed bacteriophage T4 employs one of the fastest and most powerful packaging motors, a pentamer of gp17 that translocates DNA at a rate of up to ∼2000-bp/s. We hypothesize, guided by structural and genetic analyses, that a unique hydrophobic environment in the catalytic space of gp17-Adenosine triphosphatase (ATPase) determines the rate at which the 'lytic water' molecule is activated and OH- nucleophile is generated, in turn determining the speed of the motor. We tested this hypothesis by identifying two hydrophobic amino acids, M195 and F259, in the catalytic space of gp17-ATPase that are in a position to modulate motor speed. Combinatorial mutagenesis demonstrated that hydrophobic substitutions were tolerated but polar or charged substitutions resulted in null or cold-sensitive/small-plaque phenotypes. Quantitative biochemical and single-molecule analyses showed that the mutant motors exhibited 1.8-To 2.5-fold lower rate of ATP hydrolysis, 2.5-To 4.5- fold lower DNA packaging velocity, and required an activator protein, gp16 for rapid firing of ATPases. These studies uncover a speed control mechanism that might allow selection ofmotors with optimal performance for organisms' survival.

Original languageEnglish (US)
Pages (from-to)11437-11448
Number of pages12
JournalNucleic acids research
Volume45
Issue number19
DOIs
StatePublished - Jan 1 2017

ASJC Scopus subject areas

  • Genetics

Fingerprint Dive into the research topics of 'Altering the speed of a DNA packaging motor from bacteriophage T4'. Together they form a unique fingerprint.

  • Cite this

    Lin, S., Alam, T. I., Kottadiel, V. I., Vangessel, C. J., Tang, W. C., Chemla, Y. R., & Rao, V. B. (2017). Altering the speed of a DNA packaging motor from bacteriophage T4. Nucleic acids research, 45(19), 11437-11448. https://doi.org/10.1093/nar/gkx809