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Cystine binding protein isolated from Escherichia coli W3092 in high yield was used to develop a binding assay for the disulfide amino acid cystine. Filtration of a mixture of [14C]cystine and cystine binding protein through nitrocellulose filters separated the [14C]cystine-cystine binding protein complex from the unbound cystine. The binding of [14C]cystine-cystine binding protein complexes to the filter was optimal (100%) at pH 4 to 5, and was insensitive to Mg2+ (10 mm), EDTA (10 mm), or concentrations of NaCl to 0.5 m. Bovine serum albumin in excess of 30 µg decreased binding of the 14C complex to the filter. The binding of [14C]cystine to the cystine binding protein was inhibited by lanthionine, much less by diaminopimelic acid, cystathionine, and d-cystine, and was not inhibited by reduced or oxidized glutathione, penicillamine disulfide, cysteic acid, methionine, taurine, homocystine, carboxymethyl cysteine, or the N-ethylmaleimide adduct of cysteine. Nonradioactive cystine competed with [14C]cystine resulting in the expected diminution of the bound radioactivity, permitting the direct calculation of nonradioactive cystine concentrations of unknown samples. With the cystine binding protein assay, the cystine concentrations of human plasma were in the range of 87 to 100% of the values obtained by ion exchange chromatography, and the cystine contents of cultured skin fibroblasts from patients with nephropathic cystinosis were in the range of 89 to 121% of the values obtained by ion exchange chromatography. As little as 10 pmoles of cystine were measured.

Peroxyacetyl nitrate was reactive with small molecular-weight sulfur-containing compounds The order of susceptibility was cysteine > reduced lipoic acid = reduced lipoamide > oxidized lipoic acid > oxidized lipoamide > methionine ⪢ cystine. From thiols the predominant product was disulfide. In the early stages of oxidation methionine yielded methionine sulfoxide. Products of oxidation of oxidized lipoic acid and lipoamide were the respective sulfoxides. Cystine was resistant to oxidation, yielding cysteic acid when oxidation took place. Papain was readily inactivated by peroxyacetyl nitrate while lysozyme was resistant. The small amount of inactivation of lysozyme was correlated with methionine oxidation. Papain inactivation was correlated with thiol oxidation and could be reversed by thiol compounds. The oxidation product was judged to be a dimer by methods for determining molecular weight.

S-β-(4-pyridylethyl)-l-cysteine (PEC) was transformed to S-β-(4-pyridylethyl)-l-cysteine sulfone (PEC·SO2) and to cysteine sulfinic acid following exposure to performic acid. Acid hydrolysis (6n HCl, 110°C, 20 h) of the isolated PEC·SO2 resulted in its partial decomposition to cysteic acid and alanine. Several minor ninhydrin positive derivatives, including glycine and serine, were also observed. The transformations involving PEC and PEC·SO2 were followed using high-voltage electrophoresis (HVE) at pH 1.6, 2.0, 3.5, and 6.5 and through use of the amino acid analyzer and gas-liquid chromatography. These studies indicate that as a modification form of the thiol groups of proteins, PEC is not the derivative of choice when subsequent treatment of the protein involves the use of performic acid.

The majority of the free amino acids of amniotic fluid decrease in concentration with the progression of pregnancy. Some exceptions include cysteic acid, which increases in concentration with pregnancy; taurine, methionine sulfoxide, and cystathionine, which peak around 33 to 37 weeks; ethanolamine, which is lowest at 33 to 37 weeks; and 3-methylhistidine, which is present in specimens only from 15 to 20 weeks of gestation. Published reports are summarized and salient differences are noted.

Purified pig liver monoamine oxidase, which has been shown to contain one mole of covalently bound FAD per mole of enzyme, was inhibited by [ 14 C] Pargyline (N-methyl-N-2-(propynyl)-benzylamine) and then extensively degraded by pronase. The Pargyline-containing fragment was purified by gel filtration and ion-exchange chromatography. The equimolar ratio between Pargyline and flavin was retained after the purification. Thin-layer chromatography in several systems showed that Pargyline was bound to the flavo-peptide. Amino acid analyses of the peptide yielded cysteic acid, aspartic acid, serine and glycine in a molar ratio of 1:1:1:2.

1. 1. l -Aspartate transport activity was optimally induced following 6 h growth on acetate minimal media. 2. 2. The observed aspartate (and glutamate) transport activity was attributed to the general amino acid transport system since l -phenylalanine, but not l -cysteic acid, inhibited transport activity at all pH values tested. 3. 3. Changes in K m values for l -aspartate and l -glutamate transport following growth on acetate minimal media indicated that apparent “affinities” of the “acidic amino acid transport systems” dramatically increased during the stages of active growth.

The dry and wet wrinkle-recovery of wool fabrics can be improved by reaction with metal salts in the anionic or cationic form. Small amounts of substantive metals are shown to have produced significant improvements in wet wrinkle-recovery, but, with the exception of chromium, only relatively large amounts of metals such as mercury, uranium, tungsten, and molybdenum improved dry wrinkle-recovery. The mechanism by which mercuric acetate improves dry wrinkling does not appear to involve sulphur, despite extensive reaction between mercury salts and sulphur-containing residues, such as cysteine, cystine, and cysteic acid.

Several chemical methods have been tested for their suitability as screening procedures for estimation of the protein content and quality of cowpea (Vigna unguiculata) meals. Preliminary estimates of crude protein can be obtained by the UDY dye-binding procedure, followed by the automated microKjeldahl (% N × 6.25) method using, for example, Carlo Erba equipment. Total amino acid nitrogen was estimated also with the Carlo Erba apparatus and for the cowpea varieties examined a positive correlation between the total amino acid nitrogen and total nitrogen contents was found. Total sulphur was evaluated as a measure of the sulpho-amino acids in cowpea meals. Total sulphur was determined by wet digestion and subsequent turbidimetric estimation of barium sulphate. Methionine was determined in meal hydrolysates by a Locarte Autoanalyzer. Cysteine was determined either as cysteic acid on the Locarte Autoanalyzer or as bismuth sulphide. The results obtained for cowpea meals showed that there was a relatively small variation in the total sulphur and in the sulpho-amino acid contents. Although the relation between total sulphur and sulpho-amino acids was found to be significant at the 95% confidence level, the confidence limits were wide and at this stage a definite recommendation cannot be given as to the probable usefulness of the total sulphur-screening procedure. A strong negative correlation was observed between both methionine and sulpho-amino acids/16 g of N and % crude protein. Similarly, a negative relationship was observed between total sulphur/16 g of N and % crude protein.

Three disulfide bridges out of a total of five were identified in porcine pancreatic colipase II by peptic or thermolysin digestion of the intact protein followed by separation of the cystine peptides by column chromatography and resolution of the cysteic acid fragments by diagonal electrophoresis. The position of the two other bridges could not be definitely ascertained because of the inability of the enzymes employed to cleave, to a detectable extent, the chain between the adjacent half-cystines 22 and 23. The two possible versions of the bidimensional structure of colipase suggest that the molecule is formed of two loosely bound “tails” and a heavily cross-linked central “core” probably containing the structure responsible for the colipase effect.

1. The metabolism of sulfur-bearing amino acids was investigated in R. cuneata and M. edulis, in vivo.2. Injected S35-methionine gives rise to cysteine/cystine in both animals. Cystathionine was detected in M. edulis, but not in R. cuneata though it is probably formed. A demethylation and transulfuration mechanism is postulated for this conversion.3. Both R. cuneata and M. edulis form sulfate from injected methionine. Also, both form taurine but R. cuneata does not accumulate it whereas M. edulis accumulates it in large amounts.4. Taurine is probably formed by different reactions in these species. In R. cuneata it is probably formed by decarboxylation of cysteic acid, whereas in M. edulis it is formed by oxidation of hypotaurine.

Summary Irradiation of cystine (RSSR, R = NH2CH(COOH)CH2--), cysteine (RSH), S-methycysteine (RSCH,) or lanthionine (RSR) in aqueous solution with ionizing radiation (X-or y-rays) yields products similar to those previously shown to be produced by ultraviolet light irradiation. The range and yields of products are affected by factors such as radiaion dosage, pH and the presence of air. Cystine readily yields products derived both from S-S bond fission (e.g. cysteic acid) and from C-S fission (e.g. alaninethiosulphuric acid (RSSO3H)). The initial products from cysteine are mainly cystine and alanine. With radiation doses up to 105 rad, S-methylcysteine and lanthionine usually give low yields of the corresponding sulphoxides, but with higher doses (106-107 rad) C-S fission occurs yielding other amino acids such as alanine. On irradiation of the four amino acids in the solid state, ionizing radiation, unlike 2537A light, effects only slight conversion to other amino acids. However electron spin resonance (ESR) spectra indicate that for cystine and cysteine similar trapped free radicals are obtained with both forms of electromagnetic radiation. The relative ease of S-bond fission of these amino acids on irradiation in aqueous solution as compared with solid state irradiation is compatible with accepted views on indirect action of H, OH and OOH radicals.

1. Crystalline beta-lactamase I from Bacillus cereus 569/H yielded only amino acids on acid hydrolysis, but crystalline beta-lactamase II from the same organism yielded also substantial quantities of neutral sugars and amino sugars. 2. Analysis with an amino acid analyser indicated that the two enzymes were similar though not identical in overall amino acid composition. Analysis of neutral and amino sugars as their silyl derivatives by gas-liquid chromatography showed that the carbohydrate moiety of beta-lactamase II contained residues of glucose, galactose, mannose, fucose, glucosamine and galactosamine. 3. After oxidation and hydrolysis both beta-lactamases gave small amounts of cysteic acid. After treatment of inactive Zn(2+)-free beta-lactamase II with N-ethylmaleimide or iodoacetate enzymic activity was not restored by the addition of Zn(2+).

An indirect method using glucose-U-C 14 to determine amino acid requirements was applied to fifth instar larvae of the pale western cutworm, Agrotis orthogonia Morr. C 14 O 2 was produced after glucose-U-C 14 was injected, indicating that the substrate was metabolized. The amino acids, alanine, aspartic acid, glutamic acid, proline, serine, and cysteic acid, isolated from the larvae contained relatively large quantities of radioactivity and were classed as nutritionally non-essential. Phenylalanine, arginine, lysine, histidine, leucine, iso leucine, and cystine contained little radioactivity and were classed as nutritionally essential. Methionine is probably essential also. The requirement for threonine remains in doubt. Tyrosine was not synthesized appreciably from glucose-U-C 14 but was readily formed from phenylalanine.

Cysteinesulfinic acid decarboxylase from dog liver has been purified about 300-fold. Its decarboxylating action on cysteic acid decreases during purification.

Tumors from patients with the syndrome of inappropriate secretion of antidiuretic hormone (SIADH) have been found to contain large amounts of the antidiuretic hormone vasopressin. A lung tumor from a patient with hyponatremia most likely due to SIADH was removed at surgery and found to contain 23.5 mU vasopressin/g wet weight by radioimmunoassay Slices of this tumor were incubated with phenylalanine-(3)H. Arginine vasopressin-(3)H was purified from the incubate by Sephadex G-25 column chromatography in two different systems, performic acid oxidation, and gradient elution column chromatography with diethylaminoethyl Sephadex. As oxidation of vasopressin results in drastic conformational change with breaking of the ring of the cyclic polypeptide and addition of two cysteic acid residues per molecule, the radioactive material which eluted coincident with vasopressin both before and after this procedure was considered to be arginine vasopressin-(3)H. To our knowledge this is the first demonstration of in vitro biosynthesis of vasopressin by a tumor from a patient with SIADH.Ultrastructurally, the bronchogenic carcinoma was composed of small undifferentiated and granulated cells. The granulated neoplastic cells had well developed organelles (endoplasmic reticulum, free ribosomes) concerned with protein synthesis. Secretion granules present in the tumor cells were small, surrounded by a limiting membrane, and resembled those reported in polypeptide hormone-secreting cells.

A method is described for the chromatographic separation in the Automatic Amino Acid Analyzer of the following sulfur-containing amino acids; cysteic acid, cysteinesulfinic acid, taurine, hypotaurine. The chromatographic behavior of S-sulfo-cysteine, S-sulfo-cysteamine, alaninethiosulfonic acid and thiotaurine was also studied, and methods are reported for the identification of these compounds which are not well separated from the corresponding sulfonic acids.

1. 1. Surface-active substances were isolated from the digestive juices of the crayfish Astacus leptodactylus and the lobster Homarus vulgaris, and purified by extraction procedures and chromatographic methods. 2. 2. The general structure appears to be fatty acyl-taurine and fatty acyl-dipeptide respectively. Within the dipeptide the terminal position is mainly taken in by taurine, but cysteic acid and in addition one or two unidentified amino acids also occur. The penultimate position is taken in by hydroxyproline.

HOCl by oxidative decarboxylation converts several α-amino acids into a mixture of the corresponding nitriles (major) and aldehydes (minor product). In addition, chlorination of the ring of tyrosine was observed. Cysteine when reacted with HOCl yielded cystine and cysteic acid, while with cystine, cysteic acid was the only product identified. The nitrogen bond of several dipeptides was found to be resistant to aqueous HOCl at room temperature. Chlorination of these compounds gave the corresponding N , N -dichlorodipeptide.

The reduction of the sulfoxides of methionine by Escherichia coli occurs in the presence of molecular hydrogen. By comparing the rates of reduction of some of the isomers of methionine sulfoxide it has been observed that the configuration of the asymmetric sulfur atom markedly influences this rate, whereas that of the α-carbon atom is of less significance. Under the same conditions, E. coli does not reduce l -cystine, cysteic acid, methionine sulfone, sulfanilamide, or marfanil. No evidence of racemization of the asymmetric sulfur atom by the organism was found.

The three disulfide bonds of bovine pancreatic secretory trypsin inhibitor were assigned on the basis of the structures of cystine peptides isolated from thermolysin hydrolysates of the native inhibitor prepared at pH 6.5. The peptides were isolated by chromatographic procedures with Sephadex G-75, Sephadex G-25, and Dowex 50-X2. Five major cystine peptides were oxidized with performic acid and the cysteic acid fragments were separated by chromatography on Dowex 50-X2 and by high voltage paper electrophoresis. The cysteic acid peptides were located in the amino acid sequence of the inhibitor (Greene, L. J., and Bartelt, D. C., J. Biol. Chem., 244, 2646 (1969)) on the basis of amino acid composition and end group determination. Cystine peptides corresponding to disulfide bonds I–V, II–IV, and III–VI were isolated in 55, 44, and 91% yield, respectively. Intermediates in the degradation of the inhibitor by thermolysin having either one or two peptide bonds hydrolyzed per molecule were isolated. The sites of hydrolysis were identified as the peptide bonds Ile-Leu (residues 2 to 3) and Glu-Val (residues 12 to 13).

The effect of potassium iodate on the water-soluble and acetic acid-soluble wheat proteins during mechanical dough development has been investigated. It was found that under the conditions used, cysteic acid groups were formed in the dough proteins. Performic acid-oxidised proteins when added to dough instead of potassium iodate lessen the extent of dough relaxation and produce good bread in the baking test. It is suggested that the cysteic acid groups of the proteins contribute to dough strength by forming electrostatic bonds with the basic amino-acid residues of the wheat proteins.

The amino acid sequence of the human κ Bence-Jones protein Ag containing 214 residues has been determined by a combination of four procedures: sequence analysis of the tryptic and of the chymotryptic peptides of the carboxymethylated protein described earlier and partial sequence analysis of the tryptic peptides of the aminoethylated protein and of the peptic peptides of the oxidized protein described herein. The tryptic peptides from the aminoethylated protein account for the total amino acid content and provide some necessary overlaps. Analysis of two of these peptides and of one of the peptic cysteic acid peptides has yielded the previously missing sequence for positions 19 through 32. There are two analogous disulfide bridges: one connecting Cys-23 and Cys-88 and one connecting Cys-134 and Cys-194. Comparison to the provisional sequences of other human κ Bence-Jones proteins has verified the concept that the NH2-terminal half of κ light chains is prone to variation in sequence whereas the COOH-terminal half has a fixed sequence except for one hereditary variation. Many of the differences in sequence in the NH2-terminal portion of κ-chains are not attributable to single point mutations. The various genetic mechanisms proposed to explain the variability in sequence of immunoglobulin light chains are discussed.

The synthesis of cysteitic acid from serine, phosphoserine or cysteinesulfinic acid and sulphite can be obtained in the presence of copper ions and pyruvate or pyridoxal. As previously shown, in the presence of pyridoxal, other metal ions can be substituted to the copper, but not in the presence of pyruvate. These data show that the aromatic ring of pyridoxal and its pyridine nitrogen are structural features not essential to catalytic activity in the non-enzymatic systems under study. They show also the influence of the chelated metal on the reactions of the chelate itself. The synthesis of cysteic acid from cysteinesulfinic acid and sulphite points to a new type of rupture of the HO 2 SC bond, occurring without any previous deamination of the cysteinesulfinic acid.

A number of preparations of ammonia-soluble and -insoluble fractions obtained from wool oxidized with peracetic acid has been analyzed for carbon, hydrogen, nitrogen and sulphur. Statistical examination of the results has shown that these fractions differ in carbon, hydrogen and sulphur contents, but that their nitrogen contents are identical. It is also shown by direct analysis that wool oxidized with peracetic acid does not form an ammonium salt under conditions where such a salt is formed with cysteic acid. It seems probable that strong salt links such as −SO 3 − … + NH 3 − prevent the sulphonic acid residues in oxidized wool from taking part in normal reactions.

1. Rats fed l-cystine at excessive levels excrete large amounts of sulfate sulfur in the urine and develop liver necrosis and cirrhosis. 2. Rats fed dl-methionine at excessive levels excrete large amounts of sulfate sulfur, but do not develop liver necrosis and cirrhosis. 3. Rats fed cysteic acid at high levels excrete relatively small amounts of sulfate, but do develop liver necrosis and cirrhosis. 4. In conclusion, there is no apparent correlation between the amount of sulfate formed and the development of liver damage in rats fed l-cystine, dl-methionine, and cysteic acids.

When solutions of cystine and cysteine are treated with dithiothreitol and then with sodium tetrathionate, quantitative yields of S-sulfocysteine can be obtained. This reaction has been applied to protein hydrolysates and the resulting S-sulfocysteine has been determined by ion exchange chromatography on an amino acid analyzer. With ribonuclease and reduced glutathione this procedure has given accurate values for the cystine or cysteine content of the protein or peptide. When tryptophan is also present, the addition of a small amount of sodium tetrathionate to the 6 n HCl used for the hydrolysis has usually proved to be desirable; the function of the tetrathionate appears to be to react with the indole nucleus. With this modification, the procedure has given precise results for the cysteine plus cystine contents of streptococcal proteinase, lysozyme, and wool. The results indicate that, in terms of their primary structures, cysteine and cystine are not destroyed during acid hydrolysis under these conditions; treatment with dithiothreitol and tetrathionate serves to convert any altered forms of the amino acids in the hydrolysates to S-sulfocysteine and to permit quantitative recovery in the form of this derivative. This method for determining the half-cystine content of proteins provides an alternative to the determinations of the amino acids as cysteic acid or S-carboxymethylcysteine. The treatment does not affect the recoveries of other amino acids in the hydrolysate, with the possible exception of tyrosine.

1. The polyhedral protein of silkworm virus yellows isolated by the alkali method does not contain sulfhydryl groups. 2. The content of sulfhydryl groups in the reduced polyhedral protein has been determined by titration with methylmercury nitrate and by means of Ellman's reagent (0.029–0.033 µmole of SH groups per mg of protein). The content of S-carboxymethylcysteine groups in acid hydrolysates of reduced and carboxymethylated polyhedral protein has been determined with an amino acid analyzer (0.03 µmol/mg). 3. The content of cysteic acid in acid hydrolysates of oxidized polyhedral protein is 0.069 µmole/mg.