Purification and characterization of a 28 kDa cytosolic inhibitor of cholesteryl ester hydrolases in rat testis

Lipids ◽  
1996 ◽  
Vol 31 (12) ◽  
pp. 1233-1243 ◽  
Author(s):  
Denise S. Hines ◽  
Siowfong Wee ◽  
W. McLean Grogan
Keyword(s):  
Cholesteryl Ester ◽  
Purification And Characterization ◽  
Rat Testis ◽  
Ester Hydrolases ◽  
Cytosolic Inhibitor

Purification and characterization of arginine ester hydrolases from the venom of the Chinese habu snake (Trimeresurus mucrosquamatus)

Toxicon ◽  
1984 ◽  
Vol 22 (1) ◽  
pp. 63-73 ◽  
Author(s):  
Hisayoshi Sugihara ◽  
Toshiaki Nikai ◽  
Reiko Kito ◽  
Harumi Sato
Keyword(s):  
Purification And Characterization ◽  
Ester Hydrolases

scholarly journals Purification and characterization of a fatty acid-activated protein kinase (PKN) from rat testis

1995 ◽  
Vol 310 (2) ◽  
pp. 657-664 ◽  
Author(s):  
M Kitagawa ◽  
H Mukai ◽  
H Shibata ◽  
Y Ono
Keyword(s):  
Fatty Acids ◽  
Protein Kinase ◽  
Kinase Activity ◽  
Unsaturated Fatty Acids ◽  
Limited Proteolysis ◽  
Cytosolic Fraction ◽  
Purification And Characterization ◽  
Protamine Sulphate ◽  
Rat Testis

PKN, a novel protein kinase with a catalytic domain homologous to that of the protein kinase C (PKC) family and unique N-terminal leucine-zipper-like sequences, was identified by molecular cloning from a human hippocampus cDNA library [Mukai and Ono (1994) Biochem. Biophys. Res. Commun. 199, 897-904]. Recently we partially purified recombinant PKN from COS7 cells transfected with the cDNA construct encoding human PKN, and demonstrated that the recombinant PKN was activated by unsaturated fatty acids and limited proteolysis [Mukai, Kitagawa, Shibata et al. (1994) Biochem. Biophys. Res. Commun. 204, 348-356]. The present work has focused on the further purification and characterization of PKN from native rat tissue. Immunochemical measurement revealed that PKN was found in every tissue, and was especially abundant in testis, spleen and brain; subcellular fractionation of rat brain showed that half of the PKN was localized in the soluble cytosolic fraction. PKN was purified approx. 8000-fold to apparent homogeneity from the cytosolic fraction of rat testis by DEAE-cellulose chromatography, ammonium sulphate fractionation and chromatography on butyl-Sepharose, heparin-Sepharose, Mono Q and protamine-CH-Sepharose. The enzyme migrates as a band of apparent molecular mass 120 kDa. Using serine-containing peptides based on the pseudosubstrate sequence of PKC-delta as phosphate acceptors, the kinase activity was stimulated several-fold by 40 microM unsaturated fatty acids or by detergents such as 0.04% sodium deoxycholate and 0.004% SDS. In the absence of modifiers, protamine sulphate, myelin basic protein and synthetic peptides based on the pseudosubstrate site of PKCs or ribosomal S6 protein were good substrates for phosphorylation by the kinase. In the presence of 40 microM arachidonic acid the kinase activity of PKN for these phosphate acceptors was increased 2-18-fold. The autophosphorylation activity of purified PKN was partially inhibited by pretreatment with alkaline phosphatase. These properties appear to distinguish PKN from many protein kinases isolated previously.

PURIFICATION AND CHARACTERIZATION OF DONKEY CHORIONIC GONADOTROPHIN

1980 ◽  
Vol 85 (3) ◽  
pp. 449-455 ◽  
Author(s):  
B. B. AGGARWAL ◽  
SUSAN W. FARMER ◽  
HAROLD PAPKOFF ◽  
FRANCESCA STEWART ◽  
W. R. ALLEN
Keyword(s):  
Amino Acid ◽  
Leydig Cell ◽  
Seminiferous Tubule ◽  
Chorionic Gonadotrophin ◽  
Purification And Characterization ◽  
Rat Testis ◽  
Amino Terminal ◽  
Biological Tests

Serum of the pregnant donkey, like that of the mare, contains a gonadotrophin of chorionic origin. The chorionic gonadotrophin of the donkey (dCG) has been isolated in purified form from the serum of pregnant donkeys using methodology previously employed for the purification of pregnant mare chorionic gonadotrophin (eCG). Unlike eCG, dCG is predominantly an LH in biological tests. In the in-vitro rat Leydig cell assay, dCG was as active as eCG, but in the in-vitro rat seminiferous tubule assay for FSH and in the augmentation assay, dCG was considerably less potent than eCG (1–10%). Specific rat testis radioreceptor assays for LH and FSH also showed dCG to be at least nine times more potent in LH than in FSH activity. Chemically, dCG was found to be similar to eCG in fractionation behaviour and glycoprotein nature. However, dCG had significantly less carbohydrate (31%) than had eCG (45%) and several differences were noted in a comparison of amino-acid compositions. A single amino-terminal residue, phenylalanine, was detected in dCG. Immunologically, dCG cross-reacted in homologous radioimmunoassays for eCG, equine LH and equine FSH, but its inhibition curves were all non-parallel with those of the respective equine gonadotrophin standards.

Temperature sensitivity of cholesteryl ester hydrolases in the rat testis

Lipids ◽  
1982 ◽  
Vol 17 (12) ◽  
pp. 970-975 ◽  
Author(s):  
Lucian A. Durham ◽  
W. McLean Grogan
Keyword(s):  
Cholesteryl Ester ◽  
Temperature Sensitivity ◽  
Rat Testis ◽  
Ester Hydrolases
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