Novel Human Corticosteroid-Binding Globulin Variant with Low Cortisol-Binding Affinity¹

Abstract

Corticosteroid-binding globulin (CBG) is the plasma transport protein that regulates the access of glucocorticoid hormones to target cells. Genetic deficiencies of CBG are rare, and only a single human CBG variant (Trancortin Leuven) has been related so far to decreased cortisol-binding affinity. We report here on a 43-yr-old woman, referred for chronic asthenia and hypotension, with repeatedly low morning serum cortisol levels (22-61 nmol/L; normal range, 204-546 nmol/L), normal plasma ACTH levels (38-49 pg/mL; normal, <50 pg/mL), and normal urinary cortisol (10-76 nmol/24 h; normal range, 10-105 nmol/24 h). An increased percent-free (dialysable fraction) serum cortisol (8.7-9.7%, normal range, 2.9-3.9%) suggested abnormal CBG binding activity. Indeed, she had a low serum CBG concentration (24 mg/L vs. 44+/-6 mg/L in normal women), and the affinity of her CBG for cortisol was decreased (association constant, Ka = 0.12 L/nmol vs. 0.82+/-0.29 L/nmol). In her immediate family members, the serum CBG concentration and cortisol-binding activity were normal in her husband, but the four living children had slightly lower serum CBG concentrations than the reference ranges for their pre- and postpubertal status. Measurements of cortisol distribution in undiluted serum indicated that an increase in the percentage of nonprotein-bound cortisol offsets the low cortisol levels to give approximately normal concentrations of free cortisol in serum. Direct sequencing of PCR-amplified exons encoding CBG revealed that the proband was homozygous for a polymorphism (GAC-AAC) in the codon for residue 367, which results in a Asp367-->Asn substitution. Her children were heterozygous for this polymorphism. When this nucleotide change was introduced into a normal human CBG complementary DNA, for expression in Chinese hamster ovary cells, Scatchard analysis demonstrated that the Asn367 substitution reduced the affinity of human CBG for cortisol by approximately 4-fold (Ka = 0.15 L/nmol), as compared to normal recombinant CBG (Ka = 0.66 L/nmol). These results suggest that Asp367 is an important determinant of CBG steroid-binding activity and that normal negative regulation of the hypothalamic-pituitary-adrenal axis is maintained by relatively normal serum-free cortisol concentrations, despite a marked reduction in the steroid-binding affinity of this novel human CBG variant, which we have designated as CBG-Lyon.

Structural mechanism for the carriage and release of thyroxine in the blood

“…It can now be seen that the correct site in CBG is that modeled by Edgar and Stein in 1995 (30), homologous to that shown here in TBG. Confirmation of the equivalence and validity of each of the two sites is provided by two variants of CBG with ineffective hormone binding (31,32), both involving replacements of residues, the structural equivalents of which are seen in TBG to be critical components of the hormone-binding pocket (Figs. 2a and 4a).…”

Glucocorticoid Measurements in Health and Disease - Metabolic Implications and the Potential of 24-h Urine Analyses

“…The major transport protein, CBG binds glucocorticoid hormones with high affinity thus regulating cortisol's bioavailability by restricting exit from capillaries. It is generally accepted that the CBG-bound cortisol has a restricted access to target cells [41].…”

“…The relevance of CBG as a confounder of total plasma cortisol measurements can also be deduced from the fact that CBG levels in individuals are determined, at least in part, by inheritance [41,55] and accordingly, vary significantly between individuals [43]. This can explain why those with the lowest CBG in circulation frequently also show the lowest responses to ACTH (e.g., in the short Synacthen test) which implies an increased risk of being falsely diagnosed to have an inadequate HPA reserve [43].…”

Neutrophil elastase-cleaved corticosteroid-binding globulin is absent in human plasma

“…The first immunoassays for human CBG measurements relied on the use of polyclonal antibodies and radiolabeled CBG (Robinson et al 1985). Plasma CBG concentrations measured by radioimmunoassay (RIAs) correlate well with measurements of its cortisol-binding capacity, except in samples containing CBG variants with abnormal steroid-binding properties (Robinson & Hammond 1985, Smith et al 1992, Emptoz-Bonneton et al 2000, Perogamvros et al 2010). More recently, ELISAs have been introduced that rely on the use of polyclonal antibodies as the immobilization reagent and monoclonal antibodies that recognize specific epitopes on the surface of CBG for its detection (Lewis et al 2003, Lewis & Elder 2011).…”