Biomedical subjects

W E Schmidt

Publications and source records attributed to W E Schmidt.

At least 145 records · Page 8Linked to original sources

PubMed1990

Measurement of tissue cholecystokinin (CCK) concentrations by bioassay and specific radioimmunoassay: characterization of the bioactivity of CCK-58 before and after tryptic cleavage.

Cholecystokinin (CCK) was measured in an extract of porcine brain by a bioassay system based on the enzyme release from isolated pancreatic acini and a specific radioimmunoassay. The tissue extract represented a crude peptide preparation of porcine brain that contained several molecular forms of CCK, most of them of high molecular weight including CCK-58. The different molecular forms of CCK in the porcine brain extract were isolated by high performance liquid chromatography (HPLC). Determination of CCK-like bioactivity and immunoreactivity after HPLC was performed in parallel by bioassay and radioimmunoassay, respectively CCK-58 was the most abundant molecular form in the porcine brain extract, followed by CCK-33, and CCK-8. Both assay systems measured similar relative concentrations of all different molecular forms of CCK. To elucidate the biological potency of CCK-58 before and after tryptic cleavage, a CCK-58-enriched fraction was prepared by HPLC. Part of this material was quantitatively cleaved by trypsin resulting in the formation of small molecular forms of CCK. Bioactivity and immunoreactivity of equal amounts of cleaved and uncleaved material (= CCK-58) were determined by bioassay and radioimmunoassay in parallel. Cleavage of CCK-58 increased CCK-like bioactivity by 260% and CCK-like immunoreactivity by 310%. These results indicate that in a rat pancreatic acini system, porcine CCK-58 exerts 25-30% bioactivity compared to smaller CCK forms. The specific CCK antiserum G-160 seems to possess 25-30% affinity to CCK-58 in comparison to trypsin-cleaved CCK-58. It can be concluded that this antiserum exhibits an affinity to different molecular forms of CCK which parallels their relative bioactivity.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1990

Effects of galanin, its analogues and fragments on rat isolated fundus strips.

1. Rat and porcine galanin (rGal and pGal) produced dose-dependent contraction of rat fundus strips in a concentration range of 6 nM-100 nM. 2. The stimulatory effect of rGal on rat fundus strips was not modified in the presence of somatostatin (250 nM), naloxone (1 microM), guanethidine (10 microM), a mixture of propranolol (3 microM) and phentolamine (3 microM), tetrodotoxin (1 microM), indomethacin (10 microM), atropine (1 microM), a mixture of methysergide (2.5 microM) and ketanserine (2.5 microM), a mixture of mepyramine (10 microM) and cimetidine (10 microM), and saralasin (10 microM) or when strips were desensitized to substance P and neurotensin. 3. These results suggest the localization of specific Gal receptors on the surface of smooth muscle cells of rat fundus. 4. The galanin analogues [D-Trp2]-rGal, [Nle4]-rGal, [D-Ala7]-rGal, [D-Trp2-NLe4-D-Ala7]-rGal and fragments [Cys23]-Gal (1-23), Gal (1-18) were fully active. In contrast, rGal (3-29) was completely inactive and showed no antagonistic properties to the contractile effect of intact galanin. 5. The order of potency of the galanin peptides, analogues and fragments to contract rat fundus strips was: pGal greater than rGal greater than [NLe4]-rGal greater than [Cys23]-Gal (1-23) greater than Gal (1-18) greater than [D-Ala7]-rGal greater than [Trp2]-rGal greater than [D-Trp2-NLe4-D-Ala7]-rGal. 6. The data originating from our structure-activity study suggest that the C-terminal portion of Gal contributes mainly to the affinity of Gal receptors whereas the N-terminal portion of Gal is responsible for the full activation of Gal receptors in this tissue. In particular the amino acids in position 1 and 2 of Gal (Gly-Trp) appear to be essential for binding and intrinsic activity.

Amino Acid Sequence↗

PubMed1990

Cholecystokinin receptor antagonist loxiglumide: influence on bilio-pancreatic secretion and gastrointestinal hormones in man.

We characterized the effect of the specific cholecystokinin (CCK) receptor antagonist loxiglumide (CR 1505) on gallbladder contraction, pancreatic enzyme output and plasma CCK concentrations determined by radioimmunoassay and bioassay. Gallbladder emptying and bilirubin output in response to the intraduodenal administration of a mixed liquid meal were completely inhibited by an intravenous infusion of loxiglumide (10 mg/kg/h). In contrast, meal-stimulated pancreatic enzyme secretion was diminished by only 30-40%. CCK concentrations in response to the test meal were 3-fold higher during infusion of loxiglumide, as determined by radioimmunoassay. In the absence of the antagonist, the bioassay measured CCK plasma levels identical to those determined by radioimmunoassay. In the presence of loxiglumide, CCK-like bioactivity was not detectable, indicating that the plasma concentrations of the CCK receptor antagonist were sufficient to abolish all circulating CCK-like bioactivity. We conclude that fasting volume and meal-induced contraction of the gallbladder are controlled by CCK. Postprandial pancreatic enzyme secretion, however, is mainly mediated by non-CCK-dependent mechanisms. Plasma CCK-like immunoreactivity is increased by loxiglumide, whereas plasma CCK-like bioactivity is zero in the presence of an CCK-receptor antagonist.

Adult↗

PubMed1990

Glucagon-like peptide-1(7-36)amide: characterization of the domain responsible for binding to its receptor on rat insulinoma RINm5F cells.

Glucagon-like peptide-1(7-36)amide (GLP-1(7-36)amide) is a potent stimulator of insulin secretion. Receptors for this hormone have been found on different insulinoma-derived cell lines, e.g. the RINm5F cell line which is derived from a radiation-induced rat insulinoma. To characterize the part of the GLP-1(7-36)amide molecule that is responsible for binding to its receptor on RINm5F cells, binding studies with synthetic C-terminal (GLP-1(21-36)amide) and synthetic N-terminal (GLP-1(7-25] GLP-1 fragments were carried out. GLP-1(21-36)amide showed dose-dependent binding to the GLP-1(7-36)amide receptor but was approximately 1500 times less potent in inhibiting binding of 125I-labelled GLP-1(7-36)amide than the intact hormone. GLP-1(7-25) at concentrations up to 10 mumol/l did not inhibit binding of label. Neither fragment changed intracellular cyclic AMP concentrations, in contrast to GLP-1(7-36)amide which increased intracellular cyclic AMP. GLP-1(21-36)amide, however, acted as a weak partial antagonist of GLP-1(7-36)amide with respect to GLP-1(7-36)amide-dependent stimulation of cyclic AMP production.

Animals↗

PubMed1989

CCK-antagonist L-364,718: influence on rat pancreatic growth induced by caerulein and bombesin-like peptides.

The present study investigates the inhibitory effect of the novel potent benzodiazepine-related CCK-antagonist L-364,718 on pancreatic growth in the rat induced by chronic administration of caerulein and bombesin-like peptides. Caerulein, injected s.c. twice daily at a dose of 1 microgram/kg body weight, and bombesin (10 micrograms/kg) induced a similar increase (1.5-3-fold) in pancreatic wet weight, total protein, amylase, trypsin, putrescine and spermidine content after 14 days of treatment. Growth induced by caerulein showed a significant increase in total DNA content suggesting cellular hyperplasia, whereas bombesin-like peptides led to cellular hypertrophy. In comparison to bombesin the decapeptide neuromedin C (10 micrograms/kg) was found to be 30-50% less potent. In the same dose range, neuromedin B and the tachykinins neurokinin A and B, all structurally related to bombesin, had no significant trophic effect on the rat pancreas. Administration of the CCK-antagonist L-364,718 twice daily at a dose of 0.1 mg/kg or at 1.0 mg/kg, either s.c. or orally, led dose-dependently to a near-complete inhibition of the caerulein-induced trophic effect. In contrast, L-364,718 administered at identical dosages, did not affect pancreatic hypertrophy induced by bombesin and neuromedin C. It is concluded that both peptides mediate their effect on the rat pancreas directly and not via release of endogenous cholecystokinin. Tachykinins are not involved in the regulation of pancreatic growth. Caerulein- and bombesin-like peptides have comparable effects on the stimulation of protein and polyamine synthesis.

Amino Acid Sequence↗

PubMed1989

Influence of CCK antagonist L-364,718, pancreastatin (33-49) and a somatostatin analogue on camostate-induced rat pancreatic hypertrophy.

Since endogenous cholecystokinin (CCK) is released after oral administration of camostate, it has been suggested that camostate-induced pancreatic growth is mediated via circulating CCK. To test this concept, we investigated the effects of three potentially inhibitory substances on rat pancreatic hypertrophy caused by feeding of camostate over 2 weeks: (1) L-364,718, the novel specific highly potent nonpeptide CCK receptor antagonist, (2) octreotide (SMS 201-995), a potent long-lasting somatostatin analogue and (3) pancreastatin (33-49), the biologically active C-terminal fragment of the novel gastrointestinal peptide pancreastatin. Camostate feeding (200 mg/kg) once daily for 14 days induced a significant increase in pancreatic weight, total protein, trypsinogen and polyamine levels, whereas total amylase content was substantially diminished. Simultaneous oral or subcutaneous treatment with L-364,718 (0.3 mg/kg twice daily) completely suppressed all trophic effects of camostate. Octreotide (25 micrograms/kg twice daily s.c.) and pancreastatin (33-49) (10 micrograms/kg twice daily s.c.) did not change any trophic parameter. In case of octreotide it could be shown that two daily injections only partially suppressed elevated CCK levels. Pancreatic DNA and putrescine levels were slightly reduced in rats receiving the CCK antagonist alone. These results demonstrate that camostate-induced pancreatic hypertrophy in rats is caused by the release of endogenous CCK which may contribute to the maintenance of normal pancreatic DNA and putrescine concentrations.

Animals↗

PubMed1989

Insulinotropic properties of synthetic human gastric inhibitory polypeptide in man: interactions with glucose, phenylalanine, and cholecystokinin-8.

The quantitative contribution of glucose-dependent insulinotropic polypeptide [gastric inhibitory polypeptide (GIP)] to the incretin effect after oral glucose (augmentation of insulin secretion over the degree that is explained by the glycemic rise) is not known. Therefore, hyperglycemic clamp experiments (8 mmol/L, corresponding to postprandial glucose concentrations) were performed in healthy volunteers, and synthetic human GIP was infused for 60 min at a rate (approximately 1.3 pmol/kg.min) that results in plasma GIP concentrations similar to those occurring after oral glucose loads of 75 g. The MCR for exogenous GIP was approximately 6 mL/kg.min; the decay after ceasing infusion was exponential with a t1/2 of about 18 min, and the resulting volume of distribution was about 140 mL/kg. At euglycemic (basal) plasma glucose concentrations (5.0 mmol/L) similar values were found. Insulin secretion was stimulated by hyperglycemia alone, but was greatly (2.3-fold based on C-peptide) potentiated by GIP infusions (P less than or equal to 0.001 for integrated incremental values). When integrated incremental responses over 120 min of GIP, immunoreactive insulin, and immunoreactive C-peptide were compared after oral glucose and during GIP infusions, no significant differences were found. Peak glucose concentrations after oral glucose (7.6 +/- 0.6 mmol/L) were similar to mean plasma glucose values during clamp experiments (8.2 +/- 0.1 mmol/L; P = 0.124). However, mean glucose concentrations after oral glucose were lower (6.0 +/- 0.3 mmol/L; P = 0.0004). Additional infusion of sulfated cholecystokinin-8 (25 pmol/kg.h) or the amino acid phenylalanine (1.7 mumol/kg.min) did not further stimulate insulin secretion and had no influence on the pharmacokinetics of exogenous GIP. It is concluded that human synthetic GIP is insulinotropic in man and that this activity may well explain a substantial part of the incretin effect after oral glucose. There is no interaction with cholecystokinin or phenylalanine in concentrations found after mixed meals.

Adult↗

PubMed1989

[Identification of human porins. I. Purification of a porin from human B-lymphocytes (Porin 31HL) and the topochemical proof of its expression on the plasmalemma of the progenitor cell].

We describe for the first time a porin (Porin 31HL) on the plasmalemm of an eukaryontic cell line, where porins have been found only on the outer mitochondrial membranes. The expression of the porin on the plasmalemm of transformed human B-lymphocytes is demonstrated by cytotoxicity- and indirect immunofluorescence techniques with living and fixed cells. The rabbit xenoantisera used were directed against purified Porin 31HL and free or acetylated synthetic peptides of its nineteen N-terminal amino acids. The three-step purification procedure for Porin 31HL started from a total membrane fraction of the B-cell line, followed by ion-exchange chromatography on CM- and DEAE-cellulose and a final gel filtration in SDS on Sephacryl S-300.

Amino Acids↗

PubMed1989

[Identification of human porins. II. Characterization and primary structure of a 31-lDa porin from human B lymphocytes (Porin 31HL)].

We characterize and describe for the first time the primary structure of a human porin with the molecular mass of 31 kDa derived from the plasmalemm of B-lymphocytes (Porin 31HL). Porin 31HL is shown to be a basic, channel forming membrane protein. The protein chain is composed of 282 amino acids with a relative molecular mass of 30641 Da without derivatisation. It is not a glycoprotein. The N-terminus is acetylated. Altogether the amino-acid sequence shows 56% hydrophilic or charged amino acids arranged in alternating regions of hydrophilic or hydrophobic character as it is typical for porins. In addition the 18 N-terminal amino acids of Porin 31HL can be arranged to an amphilic alpha-helix like in other porins. Porin 31HL shows approx. 29% or 24% identity to the primary structure of mitochondrial porins of Neurospora crassa and Saccharomyces cerevisiae. Partial data on mitochondrial porins from rat kidney and beef heart show sequence identity of about 90% to the human B cell porin elaborated here.

Amino Acid Sequence↗

PubMed1989

[The primary structure of crystallizable monoclonal immunoglobulin IgG1 Kol. II. Amino acid sequence of the L-chain, gamma-type, subgroup I].

The immunoglobulin Kol was the first intact antibody molecule which was characterized by high-resolution X-ray crystallography. Furthermore the complete amino-acid sequence of the heavy (H)-chain is known. Here we report the complete amino-acid sequence of the light (L)-chain of the monoclonal immunoglobulin Kol (IgG1). The polypeptide has an Mr of 22,781, consists of 216 amino acids and due to its structure is of the lambda-type. With the characteristic amino acids threonine, asparagine, threonine, glycine and lysine in positions 101, 114, 116, 154, and 165, respectively the Kol L-chain is of the Mcg isotype. With the proteins Mcg, Mot, Bur, Loc and Mem six myeloma-derived amino-acid sequences of the same isotype are known. The amino-acid sequence of the N-terminal variable part is characteristic of subgroup 1. This contribution completes the primary structure of IgG1 Kol.

Alkylation↗

PubMed1988

High-affinity binding of fungal beta-glucan fragments to soybean (Glycine max L.) microsomal fractions and protoplasts.

We have recently reported the existence of binding sites in soybean membranes for a beta-glucan fraction derived from the fungal pathogen Phytophthora megasperma f. sp. glycinea, which may play a role in the elicitor-mediated phytoalexin response of this plant [Schmidt, W. E. & Ebel, J. (1987) Proc. Natl Acad. Sci. USA 84, 4117-4121]. The specificity of beta-glucan binding to soybean membranes has now been investigated using a variety of competing polyglucans and oligoglucans of fungal origin. P. megasperma beta-glucan binding showed high apparent affinity for branched glucans with degrees of polymerization greater than 12. Binding affinity showed good correlation with elicitor activity as measured in a soybean cotyledon bioassay. Modification of the glucans at the reducing end with phenylalkylamine reagents had no effect on binding affinity. This characteristic was used to synthesize an oligoglucosyl tyramine derivative suitable for radioiodination. The 125I-glucan (15-30 Ci/mmol) provided higher sensitivity and lower detection limits for the binding assays while behaving in a manner identical to the [3H]glucan used previously. More accurate determinations of the Kd value for glucan binding indicated a higher affinity than previously shown (37 nM versus 200 nM). The 125I-glucan was used to provide the first reported evidence of specific binding of a fungal beta-glucan fraction in vivo to soybean protoplasts. The binding affinity to protoplasts proved identical to that found in microsomal fractions.

Glucans↗

PubMed1988

Purification and characterization of a 22-kDa microsomal protein from rat parotid gland which is phosphorylated following stimulation by agonists involving cAMP as second messenger.

Stimulation of secretion in exocrine glands by agonists involving cAMP as second messenger leads to the phosphorylation of the ribosomal protein S6 (protein I) and two other particulate proteins with apparent molecular masses of 24 kDa (protein II) and 22 kDa (protein III) [Jahn, R., Unger, C. & Söling, H. D. (1980) Eur. J. Biochem. 112, 345-352]. This report describes the purification and characterization of protein III. Solubilization studies indicate that protein III is an intrinsic membrane protein. It could be extracted from the endoplasmic reticulum membrane only with Triton X-100, SDS or concentrated formic or acetic acid. The purification of this protein involved extraction of the microsomes with Triton X-100, removal of the detergent by acetone precipitation, extraction of water-soluble proteins, lipids and lipoproteins, and preparative SDS polyacrylamide gel electrophoresis. The protein has a basic pI (greater than 8.7). For determination of the amino acid composition of protein III and for sequencing of its amino-terminal portion, the protein was electroeluted out off the gel, the detergent removed and the protein finally purified by reversed-phase HPLC. Protein III could be phosphorylated in vitro by the catalytic subunit of the cAMP-dependent protein kinase to a degree of approximately 0.14 mol phosphate/mol protein. The only phosphopeptide obtained after in vitro phosphorylation and subsequent tryptic or chymotryptic digestion was identical with the phosphopeptide obtained after stimulation of intact rat parotid gland lobules with isoproterenol. The sequence of this peptide was Lys-Leu-Ser(P)-Glu-Ala-Asp-Asn-Arg. It was confirmed by an analysis of the synthetic peptide following in vitro phosphorylation with cAMP-dependent protein kinase. The first 41 N-terminal residues of protein III were sequenced. So far no sequence homology with other known peptides or proteins could be found.

Amino Acids↗

PubMed1988

[Arg3]substance P and neurokinin A from chicken small intestine.

Using antisera specific for NH2-terminal and COOH-terminal regions of substance P and for the COOH-terminal region of neurokinin A, peptides with tachykinin-like immunoreactivity were isolated from extracts of chicken small intestine. The peptide Arg-Pro-Arg-Pro-Gln-Gln-Phe-Phe-Gly-Leu-Met-NH2 differs from human substance P by substitution of the lysyl residue by an arginyl residue at position 3. Synthetic [Arg3]substance P showed identical chromatographic and immunochemical properties to chicken substance P and was equipotent with substance P in contracting the guinea pig ileum. A second peptide His-Lys-Thr-Asp-Ser-Phe-Val-Gly-Leu-Met-NH2 isolated from the extracts is identical to human neurokinin A. A third peptide was immunoreactive towards the COOH-terminally directed anti-serum to substance P only but was not characterized structurally in this study.

Amino Acids↗

PubMed1988

Isolation and primary structure of tumor-derived peptides related to human pancreastatin and chromogranin A.

Using an antiserum raised against a synthetic C-terminal peptide of porcine pancreastatin, we detected pancreastatin-like immunoreactivity (PLI) in human pancreatic islets, adrenal medulla, and endocrine tumors. From a carcinoid liver metastasis, human PLI was extracted and purified by HPLC. Two C-terminally amidated peptides were isolated and characterized by sequence analysis. The first peptide, hCgA-210-301, consists of 92 amino acid residues with glycinamide as C terminus. It is identical to the cDNA-derived sequence of human chromogranin A, positions 210-301, which is preceded by two basic residues indicating a putative processing site. The C-terminal part, positions 250-301, shows 70% sequence identity to porcine pancreastatin and represents the human pancreastatin-like sequence. The second peptide, hCgA-273-301, represents a C-terminally amidated fragment of the human pancreastatin sequence, generated by an Asp-Pro cleavage at the N terminus. Peptide hCgA-273-301 was synthesized to confirm the structure of the natural peptide. Two other peptides derived from human chromogranin A were isolated and partially characterized. They are generated by proteolytic cleavage after dibasic amino acids Lys-Arg (positions 338-339) and after Trp-376 of the human chromogranin A sequence, respectively. These results indicate that chromogranin A may represent the precursor for pancreastatin-related and possibly other yet-unidentified peptides of unknown physiological function.

Amino Acid Sequence↗

PubMed1988

Pancreastatin: molecular and immunocytochemical characterization of a novel peptide in porcine and human tissues.

Pancreastatin, a novel 49-amino acid peptide isolated from porcine pancreas, shows over 70% sequence homology to the central part of bovine and human chromogranin-A. Using an N-terminal and C-terminal synthetic peptide, we developed two sensitive and specific RIAs for the detection of pancreastatin-like immunoreactivity (PLI) in porcine and human tissue extracts. PLI was present throughout the gastrointestinal tract and in most endocrine and neuronal tissues. Highest concentrations were measured in the pituitary, adrenal gland, and pancreas (1200-4000 pmol/g), similar to the distribution of chromogranin-A. PLI was also detected in human endocrine tumors, with large quantities in some carcinoids (up to 14 nmol/g). HPLC revealed that extracts from porcine pituitary and pancreas contained small pancreastatin-like peptides, whereas in adrenal medulla large chromogranin-A-like molecular forms predominated. Human endocrine tumors showed a different pattern, with intermediate forms distinct from chromogranin-A and pancreastatin. Biochemical analysis was confirmed by immunocytochemistry localizing PLI in pancreatic islets, adrenal medulla, pituitary, duodenum, and human endocrine tumors. Pancreastatin is present in a variety of gastrointestinal, endocrine, and neuronal tissues and may represent a novel peptide of unknown physiological function, derived from chromogranin-A by proteolytic cleavage.

Amino Acid Sequence↗

PubMed1987

Identification of the C-terminally alpha-amidated amino acid in peptides by high-performance liquid chromatography.

A sensitive method for the rapid identification of the C-terminally amidated amino acid in peptides is described. Peptides containing the alpha-amide group at the C-terminus were cleaved with endopeptidases. The fragments released (oligopeptides, amino acids and the C-terminally amidated residue) are coupled to phenylisothiocyanate. The phenylthiocarbamoyl derivative of the amino acid alpha-amide is selectively extracted from the mixture by alkaline butyl acetate and identified by a high-performance liquid chromatography system that enables rapid and complete separation of the derivatives of 17 amino acid amides at a detection limit of 20-50 pmol. The C-terminal alpha-amides of neurokinin-A (Met-NH2), mammalian secretin (Val-NH2), pancreatic polypeptide (Tyr-NH2) and peptide HI (Ile-NH2) are unequivocally determined at a level of 0.5-2 nmol per peptide. This method was used to characterize a crude peptide fraction prepared from porcine brain. Cholecystokinin-58 was identified in this fraction by detection of phenylthiocarbamoyl-phenylalaninamide. The method is suitable for the identification of the C-terminal alpha-amidated residue of purified peptides, but can also be used as a screening strategy to isolate from complex biological extracts novel peptides containing an alpha-amidated amino acid at the C-terminus.

Amides↗

PubMed1987

Valosin stimulates gastric and exocrine pancreatic secretion and inhibits fasting small intestinal myoelectric activity in the dog.

Valosin, a novel 25-amino acid gastrointestinal peptide with N-terminal valine and C-terminal tyrosine, has recently been isolated from porcine upper gut extracts. Its physiologic role is unknown and it does not belong to one of the structurally related gut peptide families. Assuming that valosin may influence gastrointestinal functions, we investigated the effect of high-performance liquid chromatography-pure valosin on gastric and exocrine pancreatic secretion and on the intestinal myoelectric activity in conscious dogs. Intravenous injection of valosin (0.125-1 microgram/kg) dose-dependently increased gastric acid secretion 80-fold over basal, corresponding to 18% of the maximal pentagastrin-induced effect. Pepsin output increased 10-fold over basal (30% of the pentagastrin-stimulated secretion). Half-maximal stimulation by pentagastrin could be further increased dose-dependently by simultaneous administration of valosin. Pancreatic bicarbonate secretion was stimulated 11-fold over basal at 1.0 microgram/kg, reaching about 6% of the secretin-induced maximal output, whereas protein secretion increased 12-fold over basal, corresponding to about 55% of the cholecystokinin-induced maximal output. In fasted dogs, spontaneously occurring migrating myoelectric complexes were substantially delayed during infusion of valosin at a dose of 0.2 microgram/kg. These experiments indicate that valosin may represent a novel member of the regulatory gastrointestinal peptides.

Animals↗

PubMed1987

Specific binding of a fungal glucan phytoalexin elicitor to membrane fractions from soybean Glycine max.

Treatment of soybean tissues with elicitors results in the production of phytoalexins, one of a number of inducible plant defense reactions against microbial infections. The present study uses a beta-1,3-[(3)H]glucan elicitor fraction from Phytophthora megasperma f. sp. glycinea, a fungal pathogen of soybean, to identify putative elicitor targets in soybean tissues. Use of the radiolabeled elicitor disclosed saturable high-affinity elicitor binding site(s) in membrane fractions of soybean roots. Highest binding activity is associated with a plasma membrane-enriched fraction. The apparent K(d) value for beta-glucan elicitor binding is approximately 0.2 x 10(-6) M and the maximum number of binding sites is 0.5 pmol per mg of protein. Competition studies with the [(3)H]glucan elicitor and a number of polysaccharides demonstrate that only polysaccharides of a branched beta-glucan type effectively displace the radiolabeled ligand from membrane binding. Differential displacing activity of the glucans on P. megasperma elicitor binding corresponds closely to their respective ability to elicit phytoalexin production in a cotyledon bioassay.

Journal Article↗