Cloning and Expression of Biologically Active Human Fertility Hormones

Reddy, Vemuri B.; Beck, A K; Vellucci, Vincent F.; Garramone, Anthony J.; Bernstine, Edward G.

doi:10.1007/978-1-4684-5140-5_11

Cited by 2 publications

(1 citation statement)

References 21 publications

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“…Since significantly high levels of expression could be achieved by VV-based system, low expression of hCG by other expression systems may not be due to intrinsic instability of the molecule as suggested by Reddy et al (1986).…”

Section: Discussionmentioning

confidence: 99%

High expression of human chorionic gonadotrophin beta-subunit using a synthetic vaccinia virus promoter

Gupta

Chandrasekhar²,

Pal³

et al. 2001

Journal of Molecular Endocrinology

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We have constructed a recombinant vaccinia virus to express the -subunit of human chorionic gonadotrophin ( hCG), a secretory glycoprotein that is used as an antigen for a contraceptive vaccine. The cDNA encoding the subunit was cloned under the control of a synthetic promoter that could be recognised by a vaccinia virus RNA polymerase to direct transcription. The peak expression level of hCG directed by a late synthetic promoter (Psyn) was 11·5 µg/ml, a level that was at least sixfold higher than that directed by the p7·5 early/late promoter. The expressed protein was correctly processed post-translationally such that it attained a conformation with correctly folded discontinuous epitope(s) similar to that seen in native hCG.

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Section: Discussionmentioning

confidence: 99%

High expression of human chorionic gonadotrophin beta-subunit using a synthetic vaccinia virus promoter

Gupta

Chandrasekhar²,

Pal³

et al. 2001

Journal of Molecular Endocrinology

View full text Add to dashboard Cite

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The α subunit of human chorionic gonadotropin hormone synthesized in insect cells using a baculovirus vector is biologically active

et al. 1991

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Human chorionic gonadotropin (hCG) is 8 product of the rrophoblast, With the exception of trophoblast,ic and some other cancers where oncofct,al genes are expressed, the rise in hCG in blood is indicative of pregnancy. hCG is being extensively used for induction of ovulation in women and for treatment of mate infertility. One of the demonstrated roles of hCCi is the rescue of the corpus lureurn leading co the continued secretion of progesterone, which in turn sustains the endometrium and prevents its menstrual shedding. The critical role of hCG in maintaining early pregnancy is confirmed by the ability of anti-KG antibodies, induced by active [l] or passive immunization 121, to terminate pregnancy in primates. Birth control vaccines based on ihe @'-subunit of hCG are currc:tt!y under clinical trials [3]. hCG belongs io a g:tii.ip of g!ycoprotein hormones consisting of two non-identical subunits designated CY and ,8, which sion vectors under the control of mctallothionine-1 promoter [7,8] or SV40 early promoter [O]. Conventionally purified native hCG is dissociated to obtain the subunit protein. This, however, entails recovery losses and is invariably contaminated with trace amounts of the other subunit. One of the objectives of the present work was to express the N subunit of hCG independently by recombinant DNA technology. The baculovirus expression vector system utilizing AcNPV and Sf9 cells has shown great potential for the synthesis of a variety of proteins [lo]. Correct processing of exported proteins as well as post-translational modifications, have been reported to take place in this system [ 11,121. The levels of expression (0.5-800 mg/l) are much higher than other expression systems making it an attractive alternative.We report here on the expression of the (Y subunit of hCC.i in insect cells infected with a recombinant bacuko-* -vxss c~::lin_e the cthCG, and characterization of the recombinant subunir ill terms of its immunological and biological properties. We also present evidcncc that the bulk of the cvhCG synthesized by the insect cells is not secreted into the medium, perhaps due to a 'secretory load' on the system. Finally, the implications of using a very late promoter for the synthesis of an extensively modified protein using baculovirus expression vector system are discussed. 2, MATERIALS AND METHODS Virus and cellsThe recombinant baculovirus -vAcculaCG -was constructed, characterized and plaque-purified as described 1133. AcNPV and 104Published by Elsevier Science Publishers e3. V.

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Cloning and Expression of Biologically Active Human Fertility Hormones

Cited by 2 publications

References 21 publications

High expression of human chorionic gonadotrophin beta-subunit using a synthetic vaccinia virus promoter

High expression of human chorionic gonadotrophin beta-subunit using a synthetic vaccinia virus promoter

The α subunit of human chorionic gonadotropin hormone synthesized in insect cells using a baculovirus vector is biologically active

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