Functional probing of the human glucocorticoid receptor steroid-interacting surface by site-directed mutagenesis - Gln-642 plays an important role in steroid recognition and binding

Lind, U (author)

Greenidge, P (author)

Gillner, M (author)

Koehler, K F (author)

Wright, Anthony P H (author): Karolinska Institutet,Södertörns högskola,Avdelning Naturvetenskap,Karolinska Institute

(creator_code:org_t)

2000
2000
English.
In: Journal of Biological Chemistry. - 0021-9258 .- 1083-351X. ; 275:25, s. 19041-19049

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Abstract Subject headings

To elucidate which amino acids in the glucocorticoid receptor ligand-binding domain might be involved in determining steroid binding specificity by interaction with the D-ring of glucocorticoids, we have performed site-directed mutagenesis of the four amino acids Met-560, Met-639, Gln-642, and Thr-739 based on their proximity to the steroid in a model structure. Mutations of these residues affected steroid binding affinity, specificity, and/or steroid-dependent transactivation. The results indicate that these residues are located in close proximity to the ligand and appear to play a role in steroid recognition and/or transactivating sensitivity, possibly by changes in the steroid-dependent conformational change of this region, resulting in the formation of the AF-2 site. Mutation of Gln-642 resulted in a marked decrease in affinity for steroids containing a 17 alpha-OH group. This effect was alleviated by the presence of a 16 alpha-CH3 group to a varying degree. Thr-739 appears to form a hydrogen bond with the 21-OH group of the steroid, as well as possibly forming hydrophobic interactions with the steroid, Met-EGO and Met-639 appear to form hydrophobic interactions with the D-ring of the steroid, although the nature of these interactions cannot be characterized in more detail at this point.

By the author/editor: Lind, U; Greenidge, P; Gillner, M; Koehler, K F; Wright, Anthony ...; Carlstedt-Duke, ...

About the subject

NATURAL SCIENCES: NATURAL SCIENCES; and Biological Scien ...; and Biochemistry and ...

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