Prolactin-binding components in rabbit mammary gland: characterization by partial purification and affinity labeling

Area of Science:

• Biochemistry
• Molecular Endocrinology
• Protein Chemistry

Background:

• Prolactin (PRL) is a crucial hormone regulating mammalian reproduction and metabolism.
• Understanding the molecular structure of the PRL receptor is essential for elucidating PRL signaling pathways.

Purpose of the Study:

• To investigate the molecular characteristics of the PRL receptor from rabbit mammary gland microsomes.
• To identify the molecular weight and subunit composition of the PRL receptor binding site.

Main Methods:

• Affinity purification of PRL receptors using oPRL agarose columns.
• Electrophoretic analysis (SDS-PAGE) and silver staining.
• Affinity cross-linking of microsomal receptors with iodinated ovine PRL ([125I]oPRL) using various cross-linkers.
• Estimation of molecular weights under reducing and non-reducing conditions.

Main Results:

• Affinity purification and electrophoresis revealed multiple bands, including a prominent 32,000 mol wt band.
• Affinity cross-linking identified a 59,000 mol wt hormone-receptor complex under reducing conditions.
• Calculated binding subunit molecular weight was 32,000, consistent across different cross-linking agents.
• Similar binding components were found in rabbit kidney, ovary, and adrenal microsomes, with variations in rat liver and rabbit liver microsomes.

Conclusions:

• The predominant 32,000 mol wt component represents a major binding subunit of the rabbit mammary gland PRL receptor.
• This subunit does not aggregate via disulfide (S-S) linkages.
• The PRL receptor shares binding components with other tissues and potentially with growth hormone (GH) receptors.