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Controlling bacteriophage phi29 DNA-packaging motor by addition or discharge of a peptide at N-terminus of connector protein that interacts with pRNA.

Sun J, Cai Y, Moll WD, Guo P - Nucleic Acids Res. (2006)

Bottom Line: However, the pRNA binding and virion assembly activity were greatly reduced.The DNA-packaging efficiency with dimeric pRNA was more seriously affected by the extension than with monomeric pRNA.These results reveal a potential to turn off and turn on the motor by attaching or removing, respectively, a component to outer part of the motor, and offers an approach for the inhibition of viral replication by using a drug or a small peptide targeted to motor components.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathobiology, Purdue Cancer Center and Weldon School of Biomedical Engineering, Purdue University, West Lafayette, IN 47907, USA.

ABSTRACT
Bacteriophage phi29 utilizes a motor to translocate genomic DNA into a preformed procapsid. The motor contains six pRNAs, an enzyme and one 12-subunit connector with a central channel for DNA transportation. A 20-residue peptide containing a His-tag was fused to the N-terminus of the connector protein gp10. This fusion neither interfered with procapsid assembly nor affected the morphology of the prolate-shaped procapsid. However, the pRNA binding and virion assembly activity were greatly reduced. Such decreased functions can be switched back on by the removal of the tag via protease cleavage, supporting the previous finding that the N-terminus of gp10 is essential for the pRNA binding. The DNA-packaging efficiency with dimeric pRNA was more seriously affected by the extension than with monomeric pRNA. It is speculated that the fusion of the tag generated physical hindrance to pRNA binding, with greater influence for the dimers than the monomers due to their size. These results reveal a potential to turn off and turn on the motor by attaching or removing, respectively, a component to outer part of the motor, and offers an approach for the inhibition of viral replication by using a drug or a small peptide targeted to motor components.

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Related in: MedlinePlus

Sucrose gradient (5–20%) sedimentation to detect the binding of His-procapsid with 3H-labeled pRNA (A) I-i′; (B) A-a′.
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fig4: Sucrose gradient (5–20%) sedimentation to detect the binding of His-procapsid with 3H-labeled pRNA (A) I-i′; (B) A-a′.

Mentions: The inhibition by the His-procapsid in pRNA binding was clearly shown by sucrose gradient sedimentation. The purified His-procapsid was incubated with [3H]pRNA and then subjected to 5–20% sucrose gradient sedimentation. Results showed that the binding of pRNA to His-procapsid was almost undetectable. No radioactive peak for His-procapsid–pRNA complex could be found, while a peak representing the normal procapsid–pRNA complex appeared and was centered near fraction 22 when procapsid was mixed with [3H]pRNA (Figure 4).


Controlling bacteriophage phi29 DNA-packaging motor by addition or discharge of a peptide at N-terminus of connector protein that interacts with pRNA.

Sun J, Cai Y, Moll WD, Guo P - Nucleic Acids Res. (2006)

Sucrose gradient (5–20%) sedimentation to detect the binding of His-procapsid with 3H-labeled pRNA (A) I-i′; (B) A-a′.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC1636484&req=5

fig4: Sucrose gradient (5–20%) sedimentation to detect the binding of His-procapsid with 3H-labeled pRNA (A) I-i′; (B) A-a′.
Mentions: The inhibition by the His-procapsid in pRNA binding was clearly shown by sucrose gradient sedimentation. The purified His-procapsid was incubated with [3H]pRNA and then subjected to 5–20% sucrose gradient sedimentation. Results showed that the binding of pRNA to His-procapsid was almost undetectable. No radioactive peak for His-procapsid–pRNA complex could be found, while a peak representing the normal procapsid–pRNA complex appeared and was centered near fraction 22 when procapsid was mixed with [3H]pRNA (Figure 4).

Bottom Line: However, the pRNA binding and virion assembly activity were greatly reduced.The DNA-packaging efficiency with dimeric pRNA was more seriously affected by the extension than with monomeric pRNA.These results reveal a potential to turn off and turn on the motor by attaching or removing, respectively, a component to outer part of the motor, and offers an approach for the inhibition of viral replication by using a drug or a small peptide targeted to motor components.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathobiology, Purdue Cancer Center and Weldon School of Biomedical Engineering, Purdue University, West Lafayette, IN 47907, USA.

ABSTRACT
Bacteriophage phi29 utilizes a motor to translocate genomic DNA into a preformed procapsid. The motor contains six pRNAs, an enzyme and one 12-subunit connector with a central channel for DNA transportation. A 20-residue peptide containing a His-tag was fused to the N-terminus of the connector protein gp10. This fusion neither interfered with procapsid assembly nor affected the morphology of the prolate-shaped procapsid. However, the pRNA binding and virion assembly activity were greatly reduced. Such decreased functions can be switched back on by the removal of the tag via protease cleavage, supporting the previous finding that the N-terminus of gp10 is essential for the pRNA binding. The DNA-packaging efficiency with dimeric pRNA was more seriously affected by the extension than with monomeric pRNA. It is speculated that the fusion of the tag generated physical hindrance to pRNA binding, with greater influence for the dimers than the monomers due to their size. These results reveal a potential to turn off and turn on the motor by attaching or removing, respectively, a component to outer part of the motor, and offers an approach for the inhibition of viral replication by using a drug or a small peptide targeted to motor components.

Show MeSH
Related in: MedlinePlus