. The Biological bulletin. Biology; Zoology; Biology; Marine Biology. FMRFAMIDE RECEPTORS IN SQUID 187 •o c 5 o o. (SI. [Peptide], M Figure 3. Specificity of FMRFamide receptor binding. The specific binding of nA/ ['-5I]-daYFnLRFa to SQM (50 Mg protein) was determined after incubation for h at 0°C in the presence of competing peptides at the concentrations indicated. The percentage of control specific binding has been plotted against peptide concentration. Points are means (± SEM) of triplicate determinations from single experiments, except for YGGFMRFa (duplicates from one experiment
. The Biological bulletin. Biology; Zoology; Biology; Marine Biology. FMRFAMIDE RECEPTORS IN SQUID 187 •o c 5 o o. (SI. [Peptide], M Figure 3. Specificity of FMRFamide receptor binding. The specific binding of nA/ ['-5I]-daYFnLRFa to SQM (50 Mg protein) was determined after incubation for h at 0°C in the presence of competing peptides at the concentrations indicated. The percentage of control specific binding has been plotted against peptide concentration. Points are means (± SEM) of triplicate determinations from single experiments, except for YGGFMRFa (duplicates from one experiment). were from Sigma (St. Louis, MO). The YFMRFa was from Peninsula (Belmont, CA), and FnLRFa was from Cambridge Research Biochemicals (Cambridgeshire. England). The [D-F]MRFa, F[D-M]RFa. FM[D-R]Fa. and FMR[D-F]a were from Bachem (Torrance. CA). Except for daYFnLR[TIC]amide (which we made), the remaining peptides were synthesized by Dr. B. M. Dunn (University of Florida. Gainesville. FL), purified by reverse-phase HPLC. and quantitated by amino acid analysis. Aprotinin. creatine kinase, and nucleotides (Li4)GTP[-y]S. (Li2)GTP. (Li;.)GDP, and (Na:)ATP were from Boehringer Mann- heim (Indianapolis, IN). The (Li4)Gpp[NH]p and (Na2)GMP, isobutylmethylxanthine (IBMX), and digi- tonin were from Fluka (Ronkonkoma. NY). The CHAPS and CHAPSO were from Calbio-chem (La Jolla. CA). Bestatin was from Cambridge Research Biochemicals (Wilmington. DE). Preparation of squid optic lobe membranes (SQM) Squid optic lobes (12 per preparation, each ~ g wet weight) were thawed and homogenized with 12 strokes of a motor-driven Potter-Elvehjem pestle in 24 ml ho- mogenization buffer (50 mM Hepes-NaOH, pH 150 mA/NaCl, 5 mM 2-mercaptoethanol, 3 rruUEDTA, 1 mA/EGTA) containing mTl/PMSFand 5 mMben- zamidine. The homogenate was centrifuged at 3000 rpm ( X g) for 10 min, and the pellet was rehomogenized in 24 ml homogenization buffer with inhibitors and re- centrifuged. The two supernatan
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