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

Bioactive recombinant methionyl bovine prolactin: structure-function studies using site-specific mutagenesis.

D N Luck1, P W Gout, C T Beer

  • 1Department of Biochemistry, Faculty of Medicine, University of British Columbia, Vancouver, Canada.

Molecular Endocrinology (Baltimore, Md.)
|May 1, 1989
PubMed
Summary

Researchers extracted methionyl bovine prolactin (met-bPRL) from E. coli. Its bioactivity was enhanced by thioredoxin treatment, and specific mutations revealed the first alpha-helix is crucial for prolactin

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

The implications of the 1992 Presidential election for health care reform.

JAMA·1992
Same author

[Drainage of the erectile organs in the rat].

Bulletin de l'Association des anatomistes·1992
Same author

Ranitidine, cimetidine, famotidine have no effect on post-prandial absorption of ethanol 0.8 g/kg taken after an evening meal.

Alimentary pharmacology & therapeutics·1992
Same author

CT findings in a case of exuberant cervical fibrosclerosis.

AJR. American journal of roentgenology·1992
Same author

The coding of vowel identity by patients who use the Ineraid cochlear implant.

The Journal of the Acoustical Society of America·1992
Same author

Effect of exercise on uterine activity in the patient in preterm labor.

Journal of perinatology : official journal of the California Perinatal Association·1992

Area of Science:

  • Biochemistry
  • Molecular Biology
  • Endocrinology

Background:

  • Recombinant protein production in Escherichia coli is a common method for generating hormones.
  • Bovine prolactin (bPRL) is a key lactogenic hormone, and understanding its structure-activity relationship is vital.
  • Methionyl bovine prolactin (met-bPRL) is a recombinant form produced in E. coli.

Purpose of the Study:

  • To develop a method for extracting and purifying met-bPRL from transformed E. coli.
  • To investigate the bioactivity and immunoactivity of met-bPRL and its mutants.
  • To elucidate the structural regions of bPRL critical for lactogenic activity.

Main Methods:

  • Extraction and purification of met-bPRL from transformed E. coli.
  • In vitro bioassay using Nb2 lactogen cells to measure bioactivity.

Related Experiment Videos

  • Radioimmunoassay to assess immunoactivity.
  • Site-specific mutagenesis to generate bPRL variants.

    • Main Results:

    • Extracted met-bPRL showed reactivity similar to native bPRL but lower bioactivity.
    • Thioredoxin treatment significantly increased met-bPRL bioactivity.
    • Mutants with single amino acid changes retained similar bio- and immunoactivities.
    • A double mutant (Ser62Thr/Thr65Ala) had reduced bioactivity (45%), and a Y28 deletion mutant lacked bioactivity while maintaining immunoactivity.

    Conclusions:

    • The first putative alpha-helix of bPRL is essential for both binding and mitogenic (lactogenic) activity.
    • Specific amino acid residues within this region are critical for the hormone's biological function.
    • Recombinant bPRL's bioactivity can be modulated through protein engineering and redox treatment.