Polypeptide Portion Research Articles

Delayed hypersensitivity induced by carrageenan.

Carrageenan containing a trace of polypeptide induced, without adjuvant, an intense and persistent delayed hypersensitivity in guinea pigs. This hypersensitivity was transferred by peritoneal exudate cells, and the delayed reaction was not associated with demonstrable Arthus-type skin reaction after a single-dose immunization. The roles of the polysaccharide component and of the polypeptide portion of the carrageenan in induction and elicitation are discussed.

Properties of the major glycoprotein of the beef erythrocyte membrane

The major glycoprotein of the beef erythrocyte membrane has been isolated It has a molecular weight of about 180000, of which 38% is protein and 62% is carbohydrate. The carbohydrate portion consists of hexosamine (25% of the dry weight of the glycoprotein), hexose (22%) and sialic acid (16%). The polypeptide portion is characterized by an abundance of serine and threonine residues.

Immunological Properties and Conformational Differences Detected by Tritium-Hydrogen Exchange of Clostridial Ferredoxins and Apoferredoxins

Abstract Antisera were prepared against the ferredoxins from Clostridium acidi-urici and Clostridium pasteurianum and characterized by quantitative precipitin and microcomplement fixation methods. The clostridial ferredoxins are thus the smallest naturally occurring polypeptides that have been shown to produce precipitating antibodies. The ferredoxin antibody of the immune rabbit serum is contained in the IgG fraction. The antiserum prepared against C. pasteurianum ferredoxin does not react with C. acidi-urici ferredoxin. Reconstituted ferredoxin is antigenically indistinguishable from the native ferredoxin as tested by microcomplement fixation. The antisera react much more strongly with ferredoxin than with apoferredoxin. Interpretation of this observation with respect to showing the difference in the conformation of ferredoxin and apoferredoxin is limited because the possible role of the iron-sulfur chromophore of ferredoxin as an antigenic determinant is not known. However, experiments with the tritium-hydrogen exchange technique showed conformational differences between the polypeptide portions of apoferredoxin and ferredoxin. Differences between the conformations of the polypeptide chain of the oxidized and the reduced states of C. acidi-urici ferredoxin were also shown by the tritium-hydrogen exchange technique. The reduced state was found to have fewer sites for equilibration with tritium, and thus to have a conformation more compact than the oxidized state of the iron-sulfur protein. Ferredoxin reconstituted from the fully tritiated apoferredoxin was found to retain more replaceable hydrogen atoms than the native oxidized ferredoxin.

Urinary polypeptides related to collagen synthesis

Of the total urinary hydroxyproline in normal subjects and those with skeletal disorders, between 4 and 20% was nondialyzable. In some patients with Paget's disease of bone, hyperparathyroidism with osteitis fibrosa, hyperphosphatasia, and extensive fibrous dysplasia the total urinary hydroxyproline was sufficiently high to permit purification of this polypeptide hydroxyproline by gel filtration and ion exchange chromatography. The partially purified polypeptides had molecular weights between 4500 and 10,000 and amino acid compositions and physical properties resembling those of gelatin. The polypeptide fractions also contained neutral sugar and glucosamine. These fragments had been shown to be susceptible to cleavage by purified bacterial collagenase suggesting the presence of the sequence-Pro-X-Gly-Pro-Y-. After administration of proline-(14)C to patients with Paget's disease hydroxyproline-(14)C was excreted in the urine. The hydroxyproline-(14)C specific activity reached a peak in 2-4 hr and declined rapidly. The specific activity of the polypeptide (retentate) portion was severalfold greater than that of the raw urine and diffusate. When the labeled urines were subjected to gel filtration the hydroxyproline-(14)C fractions of highest molecular weight which were eluted first from the columns had the highest specific activities. Exposure of the hydroxyproline-(14)C-containing polypeptides to bacterial collagenase rendered them dialyzable. Four patients with hyperparathyroidism and osteitis fibrosa were studied before and after removal of a parathyroid adenoma, a period of transition from a predominance of bone collagen resorption to one of relatively increased bone collagen synthesis. The total urinary hydroxyproline fell rapidly after operation whereas the ratio of the polypeptide fraction to the total rose three- to fourfold. The results of these studies suggest that the urinary polypeptides represent fragments of collagen related to collagen synthesis. Changes in the ratio of these peptides to total hydroxyproline in the urine may serve as an index of new bone formation in patients with skeletal disorders.

Purification and characterization of human chorionic gonadotropin.

A purified preparation of human chorionic gonadotropin has been obtained by techniques of alcohol precipitation, ion exchange chromatography and gel filtration. Evidence is presented that the product is homogeneous in size and amino acid composition, but heterogeneous in electrical charge and sialic acid content. The polypeptide portion of the molecule has a high proline content (12.6 moles/mole of HCG) and appears to be composed of 2 identical subunits, each with a molecular weight in the range of 25,000 to 28,000. The biological activity of the purified material is in the range of 13,700 to 18,600 IU/mg. (Endocrinology 84: 298, 1969)

Glycoprotein biosynthesis in sheep thyroid slices incubated with radioactive glucosamine and leucine. II. A study of microsomal subfractions.

Synthesis of the thyroglobulin polypeptide portion is known to precede a particle-located stepwise attachment of carbohydrates, and polysomes, contrary to microsomal membranes, do not appear to be involved in glycosylation. The labeling of sheep thyroid microsomal subfractions (where thyroglobulin is the major labeled protein) has been investigated here and we have tried to correlate protein intracellular transport with glycosylation. 1.1. Rough and smooth microsomes were prepared with Cs+ essentially as described by Dallner for liver. Chemical and electron-microscope controls pointed to a relative separation.2.2. Thyroid slices were pulse-labeled with [3H]leucine and further incubated in a chase medium, for up to 4 h. A rapid transfer of protein-bound label was found to occur from ribosomes to the deoxycholate-soluble fraction of rough microsomes (including bounding membranes and vesicle contents), but no transfer from this fraction to smooth microsomes was observed. Lack of postmicrosomal supernatants showed that secretion of protein from the endoplasmic reticulum to the colloid occurred at a show rate.3.3. When slices were incubated with [14C]glucosamine, both rough and smooth microsomes were found to be labeld. Specific radioactives of protein-bound glucosamine and sialic were higher in rough than in smooth microsomes. Results suggest that glycosylation occurs within the membranes of the whole endoplasmic reticulum rather than only within the Golgi elements.

Physical and chemical studies on glycoproteins: III. The microheterogeneity of fetuin, a fetal calf serum glycoprotein

Fetuin was isolated by alcohol-metal ion fractionation. Although the homogeneity of the preparation has been demonstrated by many conventional techniques, microheterogeneity was clearly demonstrated by resolution into eight components on starch gel electrophoresis at pH 4.2. The microheterogeneity did not result from the method of preparation because normal fetal calf serum and fetuin prepared by ammonium sulfate fractionation had the same electrophoretic pattern. Furthermore, the identical bands, characteristic of the purified fetuin, were present in sera collected from five fetal calves. Treatment with neuraminidase reduced the number of bands from eight to three, indicating that sialic acid is primarily responsible for the microheterogeneity observed. Fetuin which was denatured in 8 m urea or reduced with 2-mercaptoethanol in 8 m urea failed to show definite electrophoretic bands; the importance of tertiary structure as a contributory factor to the microheterogeneity is suggested. Purified fetuin was resolved into several fractions by chromatography on DEAE-Sephadex; each fraction contained 2–3 of the bands found on electrophoresis and reveals that the fetuin variants arise from discrete structural differences. The fractions showed similar immunochemical properties, sedimentation constants, and absorption coefficients at 278 mμ. The sialic acid, hexose and hexosamine content increased significantly according to the order of elution. The increase in carbohydrate was accompanied by a small decrease in peptide content. Amino acid analysis revealed virtually no relative difference between the fractions; minor differences were demonstrated by peptide mapping. It was concluded that the microheterogeneity results primarily from differences in the number and arrangement of the sialic acid residues at the surface of the molecule. Conformational variations of the polypeptide portion of the fetuin molecule may also play a contributing role.