Cystine, determination

Radioactive L-cystathionine (10) (765 mg.) containing 6.85 x 106 counts per minute of S35 was fed to the human cystinuric patient who had served previously as the subject in an experiment demonstrating the formation of cystine from sulfur-labeled methionine (11). The same precautions were followed with regard to human experimentation involving radioactive material as in the latter experiment. After the feeding of the cystathionine, 24-hour urine specimens were collected for 3 days and sulfur distributions were determined by the titrimetric method of Fiske (6). Cystine determinations were carried out by the procedure of Sullivan, Hess, and Howard (12). 

The Sulfur Balance for Wool. Simmonds (1956) obtained a satisfactory sulfur balance for wool when cystine was determined as cysteic acid by the method of Schram et al. (1954) and both hydriodic acid-reducible sulfur and methionine were included in the sulfur balance. A micromethod for cystine determination based on the Shinohara (1935, 1936) method, however, gave values for total sulfur in amino acids 17 % higher than the results of sulfur analysis on wool. 

Extracted with 0.2 N NaOH, 0.2 N Na S, or 50% urea in 0.2 N NaOH. Cystine determined by the Block and Bolling method (Matoltsy and Balsamo, 1955). 

Reactions of Methionine, Cysteine and Cystine. Determination of Methionine by Hydrogen Peroxide Oxidation. J. biol. Chem. 129, 48l (1939). 

Cystine determined by its catalytic effect on the iodine-azide reaction... 

Schweitz et al. purified four related toxins (Rpl, RpII, RpIII, and RpIV) from sea anemone Radianthus paumotensis (Rp) and studied their pharmacological properties (9). During the course of initial NMR studies, the reported sequence of RpII was found to have errors, and was redetermined (8). Subsequently Metrione et al. determined the sequence of RpIII as well (10). T e other two Rp toxin sequences are yet to be determined. Sequences of the Rp, and several other sea anemone toxins, are shown in Table I. We have used a two letter code to denote the species consistently and this notation differs from the earlier designations of Norton and Wiithrich groups. In our notation. As la and Ax I correspond to ATX la and AP-A, respectively. From alignment of the cystines in these sequences, it is clear that Rp toxins have three disulfide bonds, as do the other toxins. 

Figure 33 The cyclo(Adm-Cyst)3 adopts a figure-eight-like helical structure. The chiral amino acid, cystine, configuration determines the helix disposition (rigjit-handed or left-handed helix). Adamantane plays an important role as a ring size controlling agent. Taken from Ref. [163] with permission.

Besada [12] described a spectrophotometric method for determination of penicillamine by reaction with nitrite and Co(II). Penicillamine is first treated with 1 M NaN02 (to convert the amino-group into a hydroxy-group), then with 0.1 M CoCl2, and finally the absorbance of the brownish-yellow complex obtained is measured at 250 nm. The process is carried out in 50% aqueous ethanol, and the pH is adjusted to 5.4— 6.5 for maximum absorbance. The calibration graph is linear over the concentration range of 0.25-2.5 mg per 50 mL, and the mean recovery (n = 3) of added drug is 99.7%. Cystine, cysteine, methionine, and other amino adds do not interfere. 

Cysteine and cystine has been determined in seawater by a method based on cathode stripping voltammetry of the copper complex [327]. 

The amounts of single amino acids excreted in urine in the conjugated form, as determined independently by Stein and Muting, are given in Tables 1 and 2. According to Stein, glycine, glutamic acid, aspartic acid, histidine, and proline are quantitatively the most important amino acids liberated in the course of urine hydrolysis. Serine, lysine, tyrosine, cysteine and cystine, threonine, alanine, valine, phenylalanine, and leucine are... 

As early as 1905 Abderhalden (Al) isolated from the hydrolyzate of the nondiffusible fraction of human urine four amino acids, i.e., leucine, alanine, glycine, and glutamic acid, and detected two others phenylalanine and aspartic acid. Some amino acids derived from this fraction have been quantitatively determined by Albanese et al. (A3) who found in the amount of the nondiffusible fraction corresponding to one liter of urine as much as 32.8 mg tryptophan, 18.0 mg phenylalanine, 16.2 mg methionine, 15.2 mg cystine, 13.1 mg arginine, 6.7 mg histidine, and 3.9 mg tyrosine. 

The synthesis of peptides containing cystine is rarely performed starting with derivatives of cystine because of the obstacle of disulfide interchange. Disulfide interchange was discovered by F. Sanger during his attempts to determine the primary... 

Determination of iodo amino acids by HPLC with inductively coupled plasma (ICP)-MS detection had LOD 35-130 pg of I, which is about one order of magnitude lower than with UVD usually applied for these compounds175. Amino acids and peptides containing sulfur, such as cysteine, cystine, methionine and glutathione, can be determined after HPLC separation by pulsed electrochemical detection, using gold electrodes176. 

The arsenal of plant defense peptides contains members capable of binding carbohydrate residues, namely /31-4 linked A -acetyl glucosamine residues that form the biopolymer chitin. The actual mode of action remains unclear. Antifungal and antimicrobial activity has been shown in vitro. For example Ac-AMP2 is a small disulfide-rich chitin-binding peptide isolated from the seeds of Amaranthus caudatus with antimicrobial activity. It differs from Ac-AMP 1 by one additional arginine residue at the C-terminus. The structure was determined by NMR and contains a cystine knot motif. Ac-AMP2 displays a so-called hevein domain partly... 

The unambiguous formation of disulfide bonds by the synthetic strategy discussed in this section is also well suited for determining the cystine networks of newly isolated natural peptides or isomers 1231... 

Air oxidation of synthetic fully reduced co-conotoxin GVIA (9) (Scheme 4), a 27-residue peptide, 64 generated a main product that possessed the identical biological activities as the natural product whose cystine framework was determined with the synthetic compound. 65 The choice of buffer system and peptide concentration can, however, dramatically affect the folding efficiency when the DMSO-mediated oxidation method was applied. 661... 

Peptide bonds are cleaved in a nonselective, but not in a completely random manner. Based on anchimeric side-chain assistance, steric factors, and bond strains, acid-labile peptide bonds are predicted to include sites containing Asp, Glu, Ser, Thr, Asn, Gin, Gly, and ProJ22l The disulfide topologies of circulin B and cyclopsychotride, backbone-cyclized peptides with three disulfide bonds, were determined by partial hydrolysis for 5 hours.[22 Occasionally, the bond between adjacent half-cystine residues is cleaved due to the nonselective nature of the mechanism of partial acid hydrolysis.[21] By this procedure, in all cases, a complex mixture of peptide fragments is produced which requires careful chromatographic separation by RP-HPLC for subsequent analysis by mass spectrometry (see Section 6.1.6.2.7). 

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