The properties of a uterine œstradiol-receptor after limited proteolysis

Rat, R.L.; Vallet-Strouve, C.; Erdős, T.

doi:10.1016/s0300-9084(75)80025-3

Cited by 21 publications

(4 citation statements)

References 12 publications

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“…The results reported here and which are in agreement with previously published data on hormonereceptor-DNA interactions [6,8,[10][11][12]171 suggest that the method can be a useful tool for more detailed studies of the binding of hormone-receptors to DNA.…”

Section: Discussionsupporting

confidence: 92%

Estradiol‐receptor‐DNA interaction: Liquid polymer phase partition

Alberga¹,

Ferrez²,

Baulieu³

1976

FEBS Letters

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Section: Discussionsupporting

confidence: 92%

Estradiol‐receptor‐DNA interaction: Liquid polymer phase partition

Alberga¹,

Ferrez²,

Baulieu³

1976

FEBS Letters

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“…and Gorski, 1975;Chamness and McGuire, 1975;Sica et al, 1976Sica et al, , 1977Puca et al, 1976) or their requirement for fresh cytosol preparations (Puca et al, 1971(Puca et al, , 1976Sica et al, 1976Sica et al, , 1977. Trypsin has been utilized to partially cleave the calf uterine estrogen receptor (Rat et al, 1974) and to extract an insoluble estrogen receptor from chicken liver nuclei (Lebeau et al, 1973(Lebeau et al, , 1974. These latter reports suggested a disaggregation procedure that would avoid the restrictions of the former methods.…”

Section: Disaggregation and Partial Purification Of The Estrogenmentioning

confidence: 99%

“…For lamb receptor preparations, 20 yg of trypsin was added for each mg of protein, and the proteolysis maintained at 0 °C for 1 h. Rat receptor preparations required 40 yg of trypsin per mg of protein for 1 h. Trypsinization was stopped by the addition of 2.5 yg of soybean trypsin inhibitor for each yg of trypsin. While trypsin undergoes rapid autolysis in the absence of Ca2+ (TEA buffer), these conditions appear optimal for effecting receptor disaggregation while minimizing loss of binding activity (Rat et al, 1974).…”

mentioning

confidence: 99%

Characterization of trypsin-treated forms of the estrogen receptor from rat and lamb uterus

Carlson¹,

Sun²,

Katzenellenbogen³

1977

Biochemistry

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“…Vulnerability is pronounced in the N-terminal domains, while the C-terminal halves are markedly resistant. T. Erdos was the first to describe a size reduction of the estradiol receptor with retained ability for ligand binding by trypsinisation in extracts [21]. Merry Shermann, working primarily with glucocorticoid receptor, coined the term meroreceptor for the steroid-binding fragments left after digestion by added enzymes [22].…”

Section: Discussionmentioning

confidence: 99%

Surface Mapping of the Ligand‐Filled C‐Terminal Half of the Porcine Estradiol Receptor by Restricted Proteolysis

Thole¹,

Maschler²,

Jungblut³

1995

European Journal of Biochemistry

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The ligand-filled 32-kDa fragment of the porcine estradiol receptor extending from His267 to the Cterminal Ile59.5 was purified to homogeneity by adsorption to mAb 13H2. The native protein was exposed at 4 "C to a panel of proteases : thermolysin, subtilisin, pronase, elastase, ficin, bromelain, endopeptidase Lys-C, both in the dimer and the monomer state, and chymotrypsin at pH 8.2 only. The digests were analysed by SDS/PAGE/Western blotting for Coomassie staining and immunostaining. Peptides were sequenced from blots. The majority of cleavage sites in upper domain E (8 out of 11) amassed in the Leu296-Leu310 stretch. Cleavage at Leu319 was seen with subtilisin and at Tyr328 with chymotrypsin. Susceptability to enzymic proteolysis was also pronounced in Thr465 -Glu470 at the center of domain E. Three peptides, 13 kDa with thermolysin, beginning at Leu337, 6 kDa and, in low yield, 5 kDa with endopeptidase Lys-C beginning at Asp473 resp. Cys417 were only obtained from the monomer substrate. The various digests featured either 27-23-kDa peptides or mixtures of 17-13-kDa and 12-7-kDa peptides separable by SDSPAGE. All peptides with N-termini between Leu297 and Ser329 reacted with mAb 13H2. The digests showed high peaks of bound estradiol in the dimer position of 32-kDa fragment controls on density gradient centrifugation at pH 7.4. However, the property of proton-driven dissociation was only preserved in the pronase, elastase and chymotrypsin digests with peptides extending beyond the His547-ArgLeuHis550 motif. The preservation of the estradiol-binding niche in the tightly complexed peptides of domain E was also demonstrated by refilling after steroid removal. The sites exposed to proteolytic enzymes and the epitope for 13H2 attachment are in good agreement with surface probability Keywords. Estradiol receptor; steroid hormone ; domain E; restricted proteolysis ; ligand binding. plots.The hormone-binding domains E of steroid receptors are large hydrophobic segments of about 250 amino acids located in their C-terminal parts [I]. In contrast to the DNA-binding domains C, of which two-dimensional NMR and X-ray diffraction data have been reported for the estradiol [2, 31 and the glucocorticoid receptor [4.5], the structure of domain E has only been the subject of predictions. alp barrels [6], parallel p strands flanked by a helices, as in serine proteases of the subtilisin type [7], and antiparallel sheets resembling the structures found in proteins (e.g. al-antitrypsin and ovalbumin) of the serine protease-inhibitor family [8] have been discussed.We have previously shown, that the C-terminal 32-kDa half of the porcine estradiol receptor [9] extending from His267 to Ile595 [lo, 111 with a mass of 37.3 kDa can be tailored to a minimal steroid-binding core of two consecutive peptides of 17 kDa (Asn304-Lys467) and (Ser468-Lys531) 7 kDa by endopeptidase Lys-C. The Achromobacter lyticus protease used in that study [lo] is a very specific and aggressive enzyme. However, only 4 (Lys302, Lys303, Lys467 and Lys529 or Lys531) o...

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The properties of a uterine œstradiol-receptor after limited proteolysis

Cited by 21 publications

References 12 publications

Estradiol‐receptor‐DNA interaction: Liquid polymer phase partition

Estradiol‐receptor‐DNA interaction: Liquid polymer phase partition

Characterization of trypsin-treated forms of the estrogen receptor from rat and lamb uterus

Surface Mapping of the Ligand‐Filled C‐Terminal Half of the Porcine Estradiol Receptor by Restricted Proteolysis

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