Trichloroacetic acid, precipitation

Studies of canavanine interaction with the tobacco hornworm and J-. miTior also revealed the marked ability of canavanine.to inhibit whole organism incorporation of [ Hjthymidine and uridine into trichloroacetic acid-precipitated materials. When canavanine is provided simultaneously with the appropriate radio-labeled precursor, ample evidence for curtailed nucleic acid metabolism emerges but protein synthesis is unaffected (Table I, exp. I). In experiment II, canavanine is allowed to assimilate... 

Assessment of protein-associated activity with trichloroacetic acid precipitation a. Dilute a small volume of the pooled radiolabeled streptavidin with saline solution such that 50 pL of the diluted solution has 104—106 cpm. b Add 50 pL of the diluted streptavidin solution to a 12 x 75-mm glass tube, followed by 500 pL of a 0.1% BSA solution in saline, c. For precipitating the proteins, add 500 pL of 10% (w/v) TCA solution in saline. 

Goitrous hypothyroidism has rarely been described in patients taking ethionamide (393), with recovery after withdrawal (394). Ethionamide inhibits both the uptake of iodine and its incorporation into trichloroacetic acid-precipitable protein (395). 

The determination of amino acid composition has been performed in hydrolysates of adrenodoxin in 6 N-HC1 at 110° C for various periods after the removal of iron from adrenodoxin by trichloroacetic acid precipitation or by a, a -dipyridyl treatment. The tentative amino acid composition is proposed to be as shown in Table 3. Adrenodoxin contains about 100 amino acid residues with an abundance of aspartic and glutamic acids, which may account for the strong affinity of adrenodoxin for... 

We shall present our findings on the chemical nature of so-called labile sulfur in adrenodoxin. (1) The determination of dehydroalanine formation gives a negative result in measuring pyruvate after mild acid hydrolysis of trichloroacetic acid precipitates of adrenodoxin (Table 12). 

Trichloroacetic acid precipitated artificial non-heme iron protein 0.17 0.02... 

Trichloroacetic acid precipitated sample prepared from 2-mercaptoethanol and iron salt 0.02 0.02... 

Nucleic acid also was found in association with the tuberculin protein. Spiegel-Adolf and Seibert investigated the problem spectrographically, and found that the tuberculin protein ( P.P.D. ) precipitated by trichloroacetic acid exhibited an absorption band at 2650-2670 A. This band was identical with that displayed by deoxyribonucleic acid. Precipitation of the tuberculin protein by ammonium sulfate afforded a product which did not display an absorption band in the ultraviolet region of the spectrum. The substance had a lower phosphorus content than that of products obtained by trichloroacetic acid precipitation. The biological activity was not impaired in any way. 

Also, 5 -AMP has been incubated with MgS04 [14], the reaction terminated by trichloroacetic acid precipitation and inorganic phosphate determined in the supernatant by the method of Fiske and SubbaRow [163], Tartrate has been included in the assay medium in order to inhibit acid phosphatase [183]. Inorganic phosphate has been assayed by the method of King [184]. 

Wada et al. (W7) studied mucoprotein and mucoproteose fractions by horizontal paper electrophoresis in veronal buffer of pH 8.6 and in acetate buffer of pH 4.5. They processed these materials from gastric juices of normals, patients with histamine-fast anacidity, and those with gastric cancer, which were collected after insulin stimulation. They also subjected the trichloroacetic acid precipitate of gastric juice to electrophoresis, as well as the supernatant fraction remaining after acetone precipitation of the trichloroacetic filtrate of the gastric juice. 

TCA = trichloroacetic acid precipitation, followed by Biuret reaction. Further references E543, E571, EN39, EN122. 

E103 Chan, K.-M., Arriaga, C., Landt, M., Smith, C.H. and Ng, R.H. (1983). Interference by hemolysis with various methods for total calcium and its correction by trichloroacetic acid precipitation. Clin. Chem. 29, 1497-1500. 

In 1966 Kraml (K7) published a procedure for determining phospholipids on perchloric acid digests of the lipid extract. This investigator uses the extraction procedure of Bloor or the trichloroacetic acid precipitation technic of Zilversmit and Davis (Zl). Digestions are carried out on a hot plate for 15 minutes at 200°-225°C in the presence of 1.2 ml of 70% HCIO4. After cooling, the volume is made up to 5.0 ml. The final perchloric acid concentration is 14%, based on the assumption that... 

White et al. [85] also administered I125-labeled bromelain orally to rats. At various times blood was drawn and the total radioactivity, the trichloroacetic acid precipitable I125 compounds, and the molecular weight profiles of the labeled proteins were determined. They found a maximum absorption in the blood, corresponding to 270 ng/mL bromelain, one hour after bromelain administration. Approximately 40% of the radioactivity in plasma could be precipitated by 10% trichloroacetic acid. Electrophoretic analysis showed one major peak of radioactivity in plasma, with a molecular weight of 26,000-30,000 daltons. This maximum peak contained 0.003% of the administered dose of bromelain per mL plasma. 

White powder. Isoelectric point 4.6 (sheep), 7.45 (pig). Soluble in water. Chymotrypsin, trypsin and pepsin destroy the gonadotrop(h)ic action of LH. Picrolonic, flavianic. picric, and trichloroacetic acids precipitate LH with retention of its physiological activity. therap cat Gonadotropic hormone. 

Insulin in vitro increases the recovery of glucose carbon in the trichloroacetic acid-precipitable protein of epididymal adipose tissue from fasted rats (Herrera and Renold, 1960 Carruthers and Winegrad, 1961), and also increases the incorporation of glucose carbon into the ribonucleic acid fraction of adipose tissue from fasted rats (Carruthers and Winegrad, 1962). That these latter effects were observed in the presence of unlabeled glycine suggests that the bulk of the incorporation was into the pento.se moiety. When adipo.se ti.ssue from fasted rats is incubated with pyruvate- -C pyruvate carbon is isolated in the ribonucleic acid fraction however, the addition of insulin does not affect this incorporation (Carruthers and Winegrad, 1962). 

Herrera and Renold (1960) have observed that cortisol in vitro decreases the recovciy of C from glycine-i-C in the trichloroacetic acid-precipitable protein of epididymal adipose tissue (sec Section IV, H). Interest in this observation centers on the possibility that it may reflect the well-known effects of the glucocorticoids on protein metabolism. It has also been noted that cortisol in vitro decreases the glucose uptake of epididymal adipose tissue incubated in the presence of adrenaline, ACTH, or growth hormone (Renold et al., 1960b). 

During the characterization of EF-2 preparation from pyBHK cells, it was noted that there was a transfer of the [ C]-adenosine moiety from NAD to a trichloroacetic acid precipitable form during incubation in a Tris-HQ buffered reaction mixture containing no toxin, only the EF-2 preparation and [adenosine- C]-NAD [7]. The transfer of label was time dependent, reaching a maximum level at 80 to 160 min of incubation. Adding diphtheria toxin fragment A to the reaction resulted in a more rapid transfer of label to an acid precipitable form, but the maximum amount of label transferred in the two reactions was similar. The ability to transfer [ C]-adenosine from NAD to an acid precipitable form suggested that the EF-2 preparations contained a novel transferase activity. This activity was probably a contaminant in the EF-2 preparations since the majority, but not all, of the EF-2 preparations examined contained the transferase activity. 

Fig. 1. Photolabeling of fragment A in the presence of three preparations of NAD radiolabeled in different positions. Reaction mixtures containing 50 mM Tris-HCl at pH 7.2, fragment A (20 juAf), and NAD (40 tiM) radiolabeled in the nicotinamide ( ), the adenylate phosphate ( ), or the adenine (A) moiety were irradiated at 0°C under a low pressure mercury lamp (predominantly 253.7 nm). Aliquots were removed at intervals and denatured in guanidine hydrochloride, and the trichloroacetic acid-precipitable radioactivity was measured. (Reproduced from [3], with permission)...

Each sample of protein (20 tiM) was irradiated at 0°C in 50 mM Tris-HCl, pH 7.2, containing 40 yM [carbonyl- AD. After 30 min, trichloroacetic acid-precipitable radioactivity was measured (mol label incorporated/mol protein) X 100 Exotoxin A was activated by incubation with 4 M urea and 15 mM dithiothreitol, and desalted on Sephadex G-50 immediately before the experiment... 

Effect of bleomycin treatment on poIy(ADP-ribose) levels. Chicken testis cells at successive stages of spermatogenesis were incubated with bleomycin for 1 hr. Trichloroacetic acid precipitates from the different cell types were used for analysis of polymeric ADP-ribose by fluorescence methods as described by Jacobson et al. (8). The poly(ADP-ribose) content increased, relative to DNA, after bleomycin treatment in premeiotic and meiotic cells, round spermatids and in elongated spermatids. No changes were observed in mature spermatozoa (Fig. 2). 

Fig. 1. ADP-ribosylation of PC12 cell lysates by Clostridium botulinum culture supernatant. Portions of lysates were P pjADP-ribosylated without (lane 1) or with (lane 2) a preceding incubation with unlabeled NAD and toxin. Both samples contained 20 pM unlabeled NAD and 100 p ml supernatant protein. 5 pM [32p]NAD was added to the mixture in sample 1 immediately and to sample 2 after first preincubating for 30 min at 37 C. Both samples were incubated with p P]NAD for 30 min at 37 C. Culture supernatant of C. botulinum strain 17784 was partially purified by precipitation with ammonium sulfate and acid (5). PCI 2 cells (12) were pelleted by centrifugation and were lysed by freezing and thawing. Labeling was performed using ca. 50 pg of cellular protein and 5pM [32p]NAD (250,000 c.p.m.) Samples were incubated for 30 min at 37°C and were then precipitated with 10% cold trichloroacetic acid. Precipitates were dissolved, neutralized and analyzed by SDS-PAGE and autoradiography. For quantitation, bands were cut out of dried gels and counted. The uppermost radioactive band represents poly(ADP-ribose) polymerase which is labeled independently of the toxin.

An increase in protein-bound silica was found in the blood plasma (trichloroacetic acid precipitate)... 

Sonicate cellsheat supernatant 95°C/5 min and trichloroacetic acid precipitate between 5 and 10% 20 5... 

Philip Siekevitz, freshly from Greenberg s laboratory, undertook this task. By the time his postdoctoral stay drew to a close, he had developed a cell-free system from rat liver in which amino acid incorporation into a trichloroacetic acid precipitable protein fraction occurred to a small, definite extent, uncomfortably close to the noise level of the system,... 

The O antigen extracted from S. typhi with trichloroacetic acid precipitated more antibodies from anti-fS. typhi) sera than did the degraded polysaccharide extracted with acetic acid. The heterogeneity of lipopolysaccharides from strains of S, typhi isolated from carriers and patients with typhoid fever has been demonstrated by Immunoelectrophoresis. ... 

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