Heterologous expression and characterization of prolactin-like protein-A: Identification of serum binding proteins

Santanu Deb, Gary P. Hamlin, Katherine F. Roby, Simon C M Kwok, Michael J. Soares

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Abstract

In this report, we describe the heterologous expression of prolactin-like protein-A (PLP-A) in Chinese hamster ovary (CHO) cells, the characterization of recombinant PLP-A, and the identification of serum PLP-A-binding proteins. CHO cell and native placental PLP-A showed similar immunoreactive characteristics and electrophoretic mobilities. N-terminal sequencing verified the identity and purity of the recombinant PLP-A species and the site of cleavage of the signal peptide from the mature secreted PLP-A species. Recombinant PLP-A lacked activity in standardized prolactin and growth hormone in vitro bioassays. Antibodies generated to recombinant PLP-A facilitated the cellular localization of PLP-A and the identification of high molecular weight PLP-A complexes. Cross-linking analyses of radioiodinated PLP-A with serum harvested from late gestation rats indicated the presence of two major cross-linked complexes migrating under reducing conditions at 130 and 250 kDa and two minor cross-linked complexes migrating at 70 and 110 kDa. Binding of PLP-A to serum proteins was specific for PLP-A and not effectively competed by other members of the prolactin/growth hormone family. The PLP-A binding species were also found in serum from non-pregnant female and male rats.

Original languageEnglish
Pages (from-to)3298-3305
Number of pages8
JournalJournal of Biological Chemistry
Volume268
Issue number5
Publication statusPublished - 15 Feb 1993
Externally publishedYes

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Staphylococcal Protein A
Prolactin
Blood Proteins
Carrier Proteins
Cricetulus
Protein Binding
Growth Hormone
Rats
Ovary
Serum
Electrophoretic mobility
Bioassay
Protein Sorting Signals
Biological Assay

Cite this

Deb, Santanu ; Hamlin, Gary P. ; Roby, Katherine F. ; Kwok, Simon C M ; Soares, Michael J. / Heterologous expression and characterization of prolactin-like protein-A : Identification of serum binding proteins. In: Journal of Biological Chemistry. 1993 ; Vol. 268, No. 5. pp. 3298-3305.
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abstract = "In this report, we describe the heterologous expression of prolactin-like protein-A (PLP-A) in Chinese hamster ovary (CHO) cells, the characterization of recombinant PLP-A, and the identification of serum PLP-A-binding proteins. CHO cell and native placental PLP-A showed similar immunoreactive characteristics and electrophoretic mobilities. N-terminal sequencing verified the identity and purity of the recombinant PLP-A species and the site of cleavage of the signal peptide from the mature secreted PLP-A species. Recombinant PLP-A lacked activity in standardized prolactin and growth hormone in vitro bioassays. Antibodies generated to recombinant PLP-A facilitated the cellular localization of PLP-A and the identification of high molecular weight PLP-A complexes. Cross-linking analyses of radioiodinated PLP-A with serum harvested from late gestation rats indicated the presence of two major cross-linked complexes migrating under reducing conditions at 130 and 250 kDa and two minor cross-linked complexes migrating at 70 and 110 kDa. Binding of PLP-A to serum proteins was specific for PLP-A and not effectively competed by other members of the prolactin/growth hormone family. The PLP-A binding species were also found in serum from non-pregnant female and male rats.",
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Heterologous expression and characterization of prolactin-like protein-A : Identification of serum binding proteins. / Deb, Santanu; Hamlin, Gary P.; Roby, Katherine F.; Kwok, Simon C M; Soares, Michael J.

In: Journal of Biological Chemistry, Vol. 268, No. 5, 15.02.1993, p. 3298-3305.

Research output: Contribution to journalArticleResearchpeer-review

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AU - Deb, Santanu

AU - Hamlin, Gary P.

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