Binding of the pheromonal steroid, 5alpha-androst-16-en-3-one, to membrane-enriched fractions of boar and rat olfactory epithelium; preliminary evidence for binding protein being a glycoprotein.
A high degree of binding of 5alpha-[3H]-androstenone was recorded in membrane-enriched fractions of porcine olfactory tissue. The specific (i.e. high affinity, low capacity) binding had a mean Ka approximately 2x10(8)M(-1). A Hill plot of the data showed a Hill coefficient of approximately 2, possibly suggesting co-operativity of binding, with binding constants increasing from 8x10(7) to 1.6x10(9)M(-1) with increasing substrate concentration. The level of specific binding of 5alpha-[3H]-androstenone was nearly 10-fold higher than in corresponding respiratory tissue preparations and was markedly reduced in the presence of excess (approximately 1 microM) unlabelled 5alpha-androstenone. Corresponding fractions derived from rat olfactory tissue showed only 25% of the binding recorded for the pig. After incubation of 5alpha-[3H]-androstenone with solubilised olfactory cilial tissue (porcine), gel filtration and chromatography on a typical "glycoprotein" column (Concanavalin A-Sepharose B) were performed. Specific binding was recorded only in fractions corresponding to glycoproteins with Mr of approximately 70-90 kDa. In a third series of experiments, fractions containing high concentrations of cilia, some still attached to the dendritic endings (as shown by electron microscopy) were obtained by a novel method involving stripping them off the nasal epithelium. The basal adenylate cyclase (AC) activity was very significantly (P<0.01) higher in olfactory, compared with respiratory, cilia; storage at -70 degrees C for 3 weeks greatly reduced AC activity. When fresh male and female porcine olfactory cilia preparations were incubated with 5alpha-androstenone plus GTP, AC activity was increased fourfold (P<0.01). However, responses of porcine respiratory cilia were not significant statistically, neither were changes in basal levels of AC activities in rat olfactory cilia.
Kraevskaya
M A
MA
Division of Biochemistry and Molecular Biology, United Medical and Dental Schools, Guy's Hospital, London SE1 9RT, UK.
Higgins
M J P
MJ
Wilson
M T
MT
Gower
D B
DB
D000262:Adenylyl Cyclases / Q000378:metabolism; D000736:Androstenes / Q000378:metabolism*; D000818:Animals; D002845:Chromatography; D002923:Cilia / Q000201:enzymology / Q000378:metabolism; D005260:Female; D006023:Glycoproteins / Q000302:isolation & purification / Q000378:metabolism*; D008297:Male; D008854:Microscopy, Electron; D008970:Molecular Weight; D009831:Olfactory Mucosa / Q000166:cytology / Q000378:metabolism* / Q000648:ultrastructure; D011485:Protein Binding; D051381:Rats; D013552:Swine; D014316:Tritium
D000736:Androstenes; D006023:Glycoproteins; D014316:Tritium; C000214:androst-16-en-3-one; D000262:Adenylyl Cyclases