Steroid-binding serum proteins

Steroid hormones are hydrophobic molecules that convey information to diverse regions of the body by way of the bloodstream. Because of their very limited solubility in water, however, steroid hormones would precipitate 

Affinity (Kd) 2 X10 M (cortisol) 4xlO- M (testosterone) 3x10 M (cortisol) 2xlO- M (testosterone) 

Factors that increase Glucocorticoids Increased estrogens Increased thyroid 

Factors that decrease Hepatic disease Increased glucocorticoids Increased androgens (2x) 

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Table 42-8. Approximate affinities of steroids for serum-binding proteins.

Thyroid hormones and most steroid hormones are associated with carrier proteins in the serum. The carrier proteins are called, appropriately, thyroxine-binding globulin, transcortin (for cortisol), and sex-steroid-binding protein. These proteins have a high affinity (Kd 10—9— 10 8 m) for their respective hormones. They buffer the concentration of free hormone and retard hormone degradation and excretion. The carrier proteins are distinguishable from the intracellular receptors for these hormones. 

A2. Andersen, C. Y., Levels of steroid-binding proteins and steroids in human preovulatory follicle fluid and serum as predictors of success in in vitro fertilization-embryo transfer treatment. J. Clin. Endocrinol. Me lab 71, 1375-1381 (1990). 

The most frequent protein in the plasma, at around 45 g is albumin. Due to its high concentration, it plays a crucial role in maintaining the blood s colloid osmotic pressure and represents an important amino acid reserve for the body. Albumin has binding sites for apolar substances and therefore functions as a transport protein for long-chain fatty acids, bilirubin, drugs, and some steroid hormones and vitamins. In addition, serum albumin binds Ca "" and Mg "" ions. It is the only important plasma protein that is not glycosylated. 

Transcortin acts as a reservoir from which a constant supply of unbound cortisol may be provided to target cells. In addition, when serum albumin levels are low, less circulating cortisol becomes bound, which yields a greater physiological effect. Not only does protein binding control the amount of biologically active cortisol available, but it also reduces the rate at which steroids are cleared from the blood and thus limits steroid suppression of corticotrophin release from the pituitary gland. 

Liquid-liquid partition chromatography techniques based on aqueous-aqueous systems have successfully been employed in the fractionation of crude human serum, purification of steroid hormone-binding proteins from human serum, isolation of basic proteins from crude bacterial extracts, purification of immunoglobulins and monoclonal antibodies, DNA fractionations by size, topology and base sequence, as well as the isolation of soluble and ribosomal RNAs in preparative amounts from bulky mixtures [10]. Highspeed CCC using PEG-dextran system has also been employed in the separation of proteins [6]. 

Only a small fraction (1% to 2%) of imconjugated testosterone exists fireely (non-protein bound) in serum or plasma. None of the conventional assay methods, including RIA, is sufficiently sensitive to quantify the free steroid directly in a protein-free ultrafiltrate of plasma. Instead, the free steroid is estimated in plasma by adding a known amount of radiolabeled compound to the sample and allowing labeled and unlabeled compounds to reach equilibrium in their competition for the same binding sites on the proteins. Bound and free radiolabeled fractions are then separated and the ratio of free labeled to total labeled compound is determined. At equilibrium, this ratio is taken as a measure of the free testosterone fraction. An estimate of serum free testosterone can then be calculated by multiplying the free testosterone fraction by the total testosterone concentration. 

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