Salt concentrations dehydrogenase

In addition to their unusual amino acid content, halophilic proteins need high salt concentrations for maintaining their structure. The volume of halophilic proteins must be measured in these extreme multicomponent solutions. In the case of halophiles, removal of salt would lead to protein denaturation. Some examples for halophilic proteins in concentrated salt solutions are given in Table 11 cf. the values for halophilic malate dehydrogenase and halophilic glutamate dehydrogenase. 

Fig. 9. Simultaneous plots of reaction rate (relative Vmax) and activation volume (AV() for the lactate dehydrogenase system as functions of salt (KSCN) concentration. This mirror image may be explained by the similarity between Eq. (134) and Eq. (136). (By courtesy of Prof. G. N. Somero, unpublished.)...

Malate dehydrogenase from H. marismortui (AMDH) is the halophilic protein that has been studied most by solution structure methods. A molar mass of 87 kg/mol was determined for the native enzyme. It is stable at high concentrations of NaCl or KC1 and unfolds and dissociates below 2.5 M salt. Pundak and Eisenberg (1981) first measured values for the solvent interactions of AMDH and found that, in contrast to nonhalophilic globular proteins in similar conditions (Bi 0.2—0.3 g/g,B3 0.01 g/g),the halophilic protein bound... 

MTT assay is a standard colorimetric assay used to determine cytotoxicity of potential medicinal agents and other toxic materials. It is based on the reduction of the tetrazolium salt MTT by viable cells. A mitochondrial dehydrogenase enzyme is able to cleave the tetrazolium rings of the pale yellow MTT and form dark purple formazan crystals, which are largely impermeable to cell membranes resulting in the accumulation of these crystals within healthy cells. Solubilization of the cells by the addition of a detergent results in the liberation of crystals, which are solubilized. The metabolic activity of cells is directly proportional to the concentration of the created formazan product (22), whose color is quantified in a colorimetric assay. 

Fio. 22. Resolution of complex II with respect to succinate dehydrogenase by various chaotropes. Complex II was suspended in 50 mM Tris-HCl, pH 8.0. After addition of 0.6 M chaotrope the concentration of complex II was 8 mg/ml. After addition of the salts, samples were taken at the intervals indicated and assayed for succinate-Qi and succinate-PMS reductase activities. Solid lines, sucdnate-ubiquinone-2(Q) reductase activity dotted line, succinate-PMS reductase activity. Resolution temperature, 0° assay temperature, 38°. The complex II preparations used in the experiments of this and subsequent Figs. 24 and 25 had specific activities between 40 and 45 moles Qj reduced by succinate per min per mg protein. From Davis and Hatefi (169). 

NADH [6,99]. The NADH oxidase from an extremely halophilic bacterium, strain AR-1, is most active at high solute concentrations. This salt dependency, in part, reflects the dissociation of the NADH dehydrogenase from the membrane [100]. The cation requirement for oxidase activity is relatively non-specific when the cations are added as the chloride salts. NADH oxidase activity is profoundly affected by the nature of the anion. The order of decreasing effectiveness (as the Na salt) mimics the lyotropic series [101] ... 

Combined ammonium and/or potassium oxalate does not cause shrinkage of erythrocytes. However, other oxalates can cause shrinkage by drawing water into the plasma. Reduction in hematocrit may be as much as 10%, causing a reduction in the concentration of plasma constituents of 5%. As fluid is lost from the cells, an exchange of electrolytes and other constituents across the cell membrane occurs. Oxalate inhibits several enzymes, including acid and alkaline phosphatases, amylase, and lactate dehydrogenase, and may cause precipitation of calcium as the oxalate salt. 

A deficiency of ll -hydroxylase is the second most common form of CAH, with an incidence of 1 per 100,000 births, and is associated with manifestations of virilization, elevated concentrations of plasma androstenedione and DHEA-S, and hypertension. The mineralocorticoid-induced hypertension is caused by an elevation of DOC 11-deoxycortisol concentrations are markedly raised in subjects with this enzyme defect,The major distinguishing characteristic of this disorder from 21-hydroxylase deficiency, besides the elevated plasma concentrations of 11-deoxycortisol, is hypertension elicited by the salt retention caused by increased concentrations of DOC, A deficiency of 3 -hydroxysteroid dehydrogenase-isomerase has been reported and leads to an elevation in the ratio of 17a-hydroxypregnenolone to that of 17a-hydroxyprogesterone and to an increased ratio of DHEA to androstenedione. In severe forms of this rare disorder, female infants have pseudohermaphroditism, and male infants present with incomplete masculinization. Patients with this disorder usually present in early infancy with complete adrenal insufficiency including salt wasting. A late-onset form has also been reported in patients with premature pubarche with hirsutism, acne, and menstrual irregularities. The... 

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