Modular assembly of chimeric phi29 packaging RNAs that support DNA packaging

Department of Chemistry, University of Southern California, LJS-251, 840 Downey Way, Los Angeles, CA 90089-0744, USA.
Biochemical and Biophysical Research Communications (Impact Factor: 2.3). 09/2008; 372(4):589-94. DOI: 10.1016/j.bbrc.2008.05.094
Source: PubMed


The bacteriophage phi29 DNA packaging motor is a protein/RNA complex that can produce strong force to condense the linear-double-stranded DNA genome into a pre-formed protein capsid. The RNA component, called the packaging RNA (pRNA), utilizes magnesium-dependent inter-molecular base-pairing interactions to form ring-shaped complexes. The pRNA is a class of non-coding RNA, interacting with phi29 motor proteins to enable DNA packaging. Here, we report a two-piece chimeric pRNA construct that is fully competent in interacting with partner pRNA to form ring-shaped complexes, in packaging DNA via the motor, and in assembling infectious phi29 virions in vitro. This is the first example of a fully functional pRNA assembled using two non-covalently interacting fragments. The results support the notion of modular pRNA architecture in the phi29 packaging motor.

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    • "Previous results from various studies, including base deletion and mutation (Bailey et al. 1990; Reid et al. 1994c; Zhang et al. 1994, 1995b, 1997), ribonuclease probing (Reid et al. 1994a; Chen and Guo 1997), oligo targeting (Zhang et al. 1995a), competition assays to inhibit phage assembly (Trottier et al. 1996), UV crosslinking to portal protein (Garver and Guo 1997), psoralen crosslinking, and primer extension (Chen and Guo 1997), indicated two functional domains that could fold independently in pRNA. The two interlocking loops, along with the double-helical packaging domain for the binding of the motor ATPase (Lee et al. 2006), are connected through the 3WJ region in pRNA and a " three-arms around a hinge " model was proposed for pRNA function (Fang et al. 2008). Flexibility at the U72U73U74 bulge region was found to be essential for pRNA activity in gearing the DNA packaging motor (Reid et al. 1994c; Zhang et al. 1997), indicating that a conformational change may be required during DNA packaging. "
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