Characterization of two uterine proteases and their actions on the estrogen receptor
We have characterized two previously undetected proteases from the calf uterine cytosol and measured their actions on the estrogen receptor. One is an exopeptidase, purified 60-fold, that hydrolyzed amino acid (lysine-, and alanine-, or leucine-) p-nitroanilide substrates and leucyl-glycylglycine, did not hydrolyze (/sup 14/C)methemoglobin, was completely inhibited by 1 mM bestatin or puromycin (specific inhibitors of leucine aminopeptidase like enzymes), and was unable to influence the sedimentation of the 8S form of the estrogen receptor in sucrose gradients containing dilute Tris buffer. A commercial porcine leucine aminopeptidase, like the calf uterine aminopeptidase, did not convert the 8S estrogen receptor to a 4S form. Evidently, removal of the N-terminal amino acid(s) from the estrogen receptor by exopeptidase action cannot alter the sedimentation of the 8S form of the receptor, or the N-terminal amino acid(s) of the receptor is (are) unaccessible or resistant to exopeptidase activity. The second, a receptor-active protease, is an endopeptidase that did not hydrolyze any of the synthetic amide or peptide substrates tested but did possess (/sup 14/C)methemoglobin-degrading activity and the ability to convert the 8S estrogen receptor to a modified 4S form in sucrose gradients containing dilute Tris buffer. The modified 4S receptor was separable from the native receptor by DEAE-cellulose chromatography. The endopeptidase did not require Ca/sup 2 +/ for activity, and its chromatographic properties were distinctly different from a previously isolated Ca/sup 2 +/-activated protease. It was inhibited by leupeptin or dipyridyl disulfide, suggesting the presence of a thiol group that is essential for its activity.
- Research Organization:
- Univ. of Rochester School of Medicine and Dentistry, NY
- DOE Contract Number:
- AC02-76EV03490
- OSTI ID:
- 5405329
- Journal Information:
- Biochemistry; (United States), Journal Name: Biochemistry; (United States) Vol. 21:25; ISSN BICHA
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
59 BASIC BIOLOGICAL SCIENCES
ANIMALS
BODY
CARBON 14 COMPOUNDS
CATTLE
COWS
DOMESTIC ANIMALS
ENZYME ACTIVITY
ENZYMES
ESTROGENS
FEMALE GENITALS
HORMONES
HYDROLASES
ISOTOPE APPLICATIONS
LABELLED COMPOUNDS
MAMMALS
ORGANS
PEPTIDE HYDROLASES
RECEPTORS
RUMINANTS
STEROID HORMONES
TRACER TECHNIQUES
UTERUS
VERTEBRATES