Jove
Visualize
Contact Us
JoVE
x logofacebook logolinkedin logoyoutube logo
ABOUT JoVE
OverviewLeadershipBlogJoVE Help Center
AUTHORS
Publishing ProcessEditorial BoardScope & PoliciesPeer ReviewFAQSubmit
LIBRARIANS
TestimonialsSubscriptionsAccessResourcesLibrary Advisory BoardFAQ
RESEARCH
JoVE JournalMethods CollectionsJoVE Encyclopedia of ExperimentsArchive
EDUCATION
JoVE CoreJoVE BusinessJoVE Science EducationJoVE Lab ManualFaculty Resource CenterFaculty Site
Terms & Conditions of Use
Privacy Policy
Policies

Related Experiment Videos

Cauliflower mosaic virus reverse transcriptase. Activation by proteolytic processing and functional alteration by

H Takatsuji1, H Yamauchi, S Watanabe

  • 1Laboratory of Molecular Genetics, National Institute of Agrobiological Resources, Ibaraki, Japan.

The Journal of Biological Chemistry
|June 5, 1992
PubMed
Summary

Related Concept Videos

You might also read

Related Articles

Articles linked to this work by shared authors, journal, and citation graph.

Sort by
Same author

Adenosine A2 receptor antagonist facilitates the reversal of long-term potentiation (depotentiation) of evoked postsynaptic potentials but inhibits that of population spikes in hippocampal CA1 neurons.

Neuroscience letters·1992
Same author

The helix-loop-helix/leucine repeat transcription factor USF can be functionally regulated in a redox-dependent manner.

The Journal of biological chemistry·1992
Same author

Immunohistochemical study of the histogenesis of esophageal carcinosarcoma.

Japanese journal of clinical oncology·1992
Same author

Structure and distribution of the lymphatic vessels in the parietal pleura of the monkey as studied by enzyme-histochemistry and by light and electron microscopy.

Archives of histology and cytology·1992
Same author

Specific expressions of Fyn and Lyn, lymphocyte antigen receptor-associated tyrosine kinases, in the central nervous system.

Brain research. Molecular brain research·1992
Same author

A hyperpolarizing response induced by glutamate in mouse cerebellar Purkinje cells.

Neuroscience research·1992

Cauliflower mosaic virus reverse transcriptase (RTase) is synthesized as an inactive precursor and activated through proteolytic processing. N-terminal deletions yield active RTase, suggesting this mechanism is crucial for enzyme function.

Area of Science:

  • Molecular Biology
  • Enzymology
  • Virology

Background:

  • Cauliflower mosaic virus (CaMV) reverse transcriptase (RTase) gene was previously expressed in yeast (RTase-Y) and E. coli (RTase-E).
  • RTase-Y (60 kDa) is active, while RTase-E (78 kDa) is inactive, indicating post-translational modification or processing.

Purpose of the Study:

  • To investigate the role of proteolytic processing in the activation of CaMV RTase during de novo synthesis.
  • To identify the specific regions of the CaMV RTase involved in its activation and enzymatic activity.

Main Methods:

  • Construction and expression of truncated CaMV RTase genes (deletions at N-terminus, C-terminus, or both) in E. coli.
  • Activity gel analysis to determine molecular size and enzymatic activity of truncated RTases.

Related Experiment Videos

  • RNA-dependent DNA synthesis assays using synthetic and natural RNA templates.
  • Characterization of metal ion requirements and substrate incorporation (dTTP).

    • Main Results:

    • N-terminal deletions (delta N) resulted in active RTases, similar in properties to yeast-expressed RTase-Y.
    • C-terminal deletions (delta C) and double deletions (delta NC) showed activity on synthetic templates but not natural RNA.
    • Delta N RTases transcribed natural RNA and required Mg2+ or Mn2+, while delta C RTases lost activity on natural RNA, potentially due to dTTP incorporation issues.

    Conclusions:

    • CaMV RTase is synthesized as an inactive precursor and activated by proteolytic processing, likely involving the N-terminal region.
    • The C-terminal region is important for activity on natural RNA templates and proper metal ion cofactor utilization.
    • Proteolytic processing is a key step in generating functional CaMV RTase from its de novo synthesized precursor.