Low Molecular Peptides Research Articles

Inhibition of exogenous mRNA translation in a cell-free system by chromatin peptides from calf thymus.

Low molecular weight chromatin peptides isolated from calf thymus by affinity chromatography on DNA-cellulose inhibit significantly translation of exogenous isolated mRNA in a reticulocyte cell-free system. Translation with endogenous mRNA present in the system is not inhibited by low peptide concentrations. The data obtained combined with the previous findings suggest that chromatin peptides control gene expression at two levels: transcription and translation.

Structure of tetanus toxin. Demonstration and separation of a specific enzyme converting intracellular tetanus toxin to the extracellular form.

Protease activity has been demonstrated in culture supernatants of Clostridium tetani at various stages of fermentation. Gel chromatography of the concentrated filtrates revealed the presence of three enzymatically active fractions eluting at separate positions off the column. The smallest protease was found to "nick" the single chain intracellular tetanus toxin, producing the extracellular, two-chain structure of the molecule. As little as 3 ng of active protease were sufficient to cleave 50 microgram of intracellular tetanus toxin, suggesting that this enzyme is responsible for the observed structural change of the toxin molecule during its release into the culture medium. By comparison, the second protease, eluting at an intermediate position, exhibited only marginal activity towards intracellular toxin. The third, largest, enzyme was not active under the conditions of the assay. However, the latter protease effectively hydrolyzed low molecular weight histidyl peptides, and it is concluded that this enzyme is similar to the one described by Miller, P.A. Gray, C.T., and Eaton, M.D. (1960) J. Bacteriol. 79, 95-102. The properties of the partially purified enzymes, including their differential behavior towards a number of protease inhibitors, are reported.

Biological Alterations Resulting from Chronic Lung Irradiation: II. Connective Tissue Alterations Following Inhalation of 144 Ce Fused Clay Aerosol in Beagle Dogs

Beagle dogs were exposed by inhalation to an aerosol of ${}^{144}{\rm Ce}$ clay to quantitate the relationship between pulmonary radiation dose and induced fibrosis. Collagen, elastin, glucosamine and the ratios of elastin/collagen, hydroxyproline/hydroxylysine and hydroxyproline/proline were determined to indicate changes in connective tissue constituents. Total lung collagen was partitioned into native collagen, soluble collagen and ultrafilterable hydroxyproline peptides. Increased total lung collagen correlated best with increasing cumulative radiation dose and increasing time after inhalation exposure. The increase in total lung collagen was not seen until more than 4 mo after exposure and a cumulative dose of about 40,000 rad. Soluble collagen and low molecular weight hydroxyproline peptide quantities both increased at 2 mo after exposure and cumulative doses of 20,000-27,000 rad. A variable elastin response apparently was not related to either increasing time or increasing radi...

Studies on the mechanism of covalent incorporation of a lysergyl derivative to immunoglobulin peptides in vitro.

In vitro incubation of hyperimmune rabbit lymphoid cells with the hallucinogenic indole alkaloid, d-lysergic acid diethylamide (LSD), results in biosynthesis of modified, secretable immunoglobulin peptides. Modification involves covalent attachment of the lysergyl moiety to COOH-terminal portions of the peptides. Aalogous effects occur when cells are incubated in vitro in the presence of the non-hallucinogenic LSD analogue, d-lysergic acid, and N-[3H]-carboxymethyl-d-lysergamide. The phenomenon is reversed by tryptophan and is inhibited by puromycin and cycloheximide. In vitro attachment of the lysergyl moiety occurs in the presence of actinomycin D at levels which inhibit RNA synthesis. While LSD is not attached to intracellular tRNA, the drug binds to 80 S ribosomes from hyperimmune lymphoid cells with high affinity (K A equals 3.5 times 10-8 M-1). Similar binding occurs to nonimmune splenic ribosomes. Implications of these findings are discussed with respect to the degree of involvement of cellular protein translational mechanisms in the covalent attachment of the lysergyl moiety to low molecular weight immunoglobulin peptides.

The influence of alkali and heat treatment on yeast protein.

AbstractThe soluble components in disintegrated cells of Saccharomyces cereivisiae have been characterized by means of extraction, centrifugation, dialysis, and gel filtration. The influence of alkali and heat treatment on the protein and RNA in the soluble fraction from disintegrated yeast cells and on functional properties of protein concentrates have been studied.After water extraction and centrifugation at 100000 g 42% of the nitrogen containing components of the disintegrated cells were recovered in the supernatant. By extraction at pH 11.5 an additional 31% of the nitrogen was solubilized. Half of the water‐soluble nitrogen‐containing components has a molecular weight lower than 5000. In the water‐ and alkali‐soluble fractions about 80% of each amino acid was recoveredThe water‐soluble protein was separated into 3 fractions by gel filtration on Sephadex G 200. The major portion of the protein had a molecular weight about 100,000. The amount of protein in this fraction was decreased after treatment at increasing pH and temperature. No degradation of protein to low molecular peptides occurred. The amount of RNA in the soluble fraction was only slightly influenced by alkali treatment and by heat treatment at pH 7.5 in the presence of 5% NaCl. RNA was not degraded to low molecular components of the treatments.The solubility of protein concentrates decreased after treatment at alkaline pH and after heat precipitation.

Ca 2+ binding study on some low molecular weight elastin peptides

The binding of Ca 2+ to some low molecular weight peptides obtained from the alkaline hydrolysis of purified elastin has been determined by titration using murexide as an indicator for free Ca 2+. Ca 2+ binding was shown to be dependent on pH and Ca 2+ concentration and increases with increasing pH. The data strongly support the possibility that electrostatic interaction occurs between the ionized carboxylic groups of the peptide and Ca 2+. In fact the peptide in which the carboxylic groups were previously blocked with the glycine ethyl ester did not interact with Ca 2+.

Enzymes lytic against Pseudomonas aeruginosa produced by Bacillus subtilis YT-25.

A bacterium YT–25 which produces enzymes lytic against Pseudomonas aeruginosa was isolated from soil and it was identified as Bacillus subtilis.A1-enzyme, A2-enzyme, B-enzyme and NLF (Native Cell-Lytic Factor) which contribute the lysis of P. aeruginosa were purified from the culture filtrate of strain YT–25.Purified A1-enzyme, A2-enzyme and B-enzyme individually lysed the vegetative cells of P. aeruginosa in the presence of NLF.NLF is a low molecular basic peptide and seemed to alter the sufrace structure of P. aeruginosa.B-enzyme hydrolyzed the peptidoglycan purified from P. aeruginosa to release the reducing groups, but A1-enzyme and A2-enzyme released neither reducing groups nor free amino groups from the peptidoglycan.

Cleavage of bacterial flagellin with proteolytic enzymes: II. Induction of enhanced delayed-type hypersensitivity to flagellin by peptic fragments of the protein

Digestion of Salmonella adelaide flagellin (mol wt 40,000) with pepsin at suboptimal pH results in the release of 3–4 large, “pepsin-resistant peptides” (mol wt 13,000–15,000), which carry most of the antigenic determinants of the protein. These large peptides are slowly degraded by prolonged pepsin treatment into low molecular weight peptic peptides (mol wt < 4,000) which have little or no antigenic activity. A single injection into rats of the peptic digests of flagellin failed to produce detectable anti-flagellin antibodies, but was still capable of priming rats for a secondary antibody response to flagellin. This priming effect was mediated by the “pepsin-resistant peptides” present in the peptic digests, but not by the low molecular weight peptic peptides. In contrast to the antibody response, preparations of flagellin that had been subjected to prolonged peptic digestion induced higher levels of flagellin-specific delayed-type hypersensitivity than undigested flagellin. Furthermore, on fractionation of such digests, it was found that both the “pepsin-resistant peptides” and the low molecular weight peptic peptides induced delayed-type hypersensitivity to flagellin. A partial purification of the small peptic peptides which provoked delayed hypersensitivity was achieved by high voltage paper electrophoresis. The hypersensitivity inducing peptides were shown to be derived from regions of the flagellin molecule which carried antibody forming determinants. From the observation that degraded antigen preferentially induces delayed hypersensitivity, it is proposed that macrophage-degraded antigen plays an important role in regulating humoral and cell-mediated immunity in vivo. It is also suggested that degraded antigen favours the induction of delayed hypersensitivity due to its inability to promote cell-cell interaction between immunocompetent cells.

Inactivation of an alkaline bacillopeptidase by pentasodium triphosphate.

AbstractThe inactivation of an alkaline microbial protease by pentasodium triphosphate (TPP) follows first order kinetics, the apparent activation energy being the same from pH 7–10 with and without added TPP. Almost exclusively low molecular peptides were observed as reaction products.

Biochemistry and Biology of a Leucotactic Binary Serum Peptide System Related to Anaphylatoxin

The purification and crystallization of the peptide components of a newly detected binary peptide system in mammalian serum (hog, rat and guinea pig) is described. The peptide system consists of two components, classical anaphylatoxin and cocytotaxin. Cocytotaxin represents a hitherto unknown basic peptide of mammalian serum which has been modified by a contact reaction with immune complexes or other hydrophilic, insoluble substances of high molecular weight. The molecular homogeneity of both crystallized peptides has been established. Classical anaphylatoxin has a molecular weight of 9,500, and cocytotaxin, which is highly similar in its physicochemical behaviour to anaphylatoxin, has a molecular weight of 8,500. Products of an enzymatic reaction with anaphylatoxin and cocytotaxin as substrates have been isolated and characterized as three proteins with molecular weights of 28,000, 56,000 and 112,000. The biological activities of both low molecular peptides were investigated. Anaphylatoxin is a well-known histamine liberator in guinea pigs and elicits anaphylactic reactions. Cocytotaxin, although physicochemically similar to anaphylatoxin, produces none of the actions of anaphylatoxin and has no known separate biological activity. Neither anaphylatoxin nor cocytotaxin display significant chemotactic activity for neutrophils. On recombination and depending on their molar ratio and the absolute concentration of each of the peptides, cell-specific chemotactic activity for neutrophil and eosinophil leucocytes is observed. Cell-specific leucotactic and anaphylatoxin activities could be demonstrated to be different activity phases of the binary peptide system with only a partial overlap. The mechanism of action of this peptide system on cells was evaluated. It was found that cocytotaxin can be replaced by nucleotides such as ATP, cyclic AMP etc., suggesting that cocytotaxin acts as regulatory peptide and that the energy metabolism is activated by anaphylatoxin as catalytic peptide. Considering the pharmacological activities of the three metabolites of the peptides, a relationship between chronic inflammation and circulatory disease may be assumed.

Disulfide interchange reactions between cystamine and cystine peptides

A method is presented for carrying out interchange reactions on a small scale between cystamine and low molecular weight cystine peptides. The products of the reaction were separated from unchanged peptides and from excess cystamine by high-voltage electrophoresis on paper. The opening of intrachain disulfide bonds required substantially higher concentrations of cystamine than did the reaction of interchain bonds. The interchange reaction can be used to separate cysteine peptides on an analytical or a preparative scale from other components of enzymic digests of proteins. A cys-gly bond in one interchange product was hydrolyzed by trypsin, but a cys-leu bond in another interchanged peptide was insensitive to trypsin.

Structure of Plasteins

THE molecular weight of the ‘plastein' precipitated from a pepsin hydrolysate of egg albumen at pH. 4 was found to be about 300 in determinations performed cryoscopically in a phenol solution1. Since the average molecular weight of peptides in pepsin hydrolysate is between 300 and 400 2, no lengthening of peptide chains seems to take place in the formation of plastein, still less protein synthesis, as had generally been thought. For this reason we regarded the plasteins as being constituted of mixtures of cyclic peptides which are formed under the influence of certain proteolytic enzymes from low-molecular peptides in protein hydrolysates1. Of the earlier investigators, Folley3 alone has supported the view of the low molecular-weight structure of plasteins.