Number Of SH Groups Research Articles

Thiol group reactivity and polymerization of actin in the presence of ATP analogs

We have investigated polymerization and the number of SH-groups of monomeric actin exposed in the presence of (beta, gamma)-substituted ATP-analogs. Actin, when depolymerized in a buffer containing 10 equiv. of APPCP exposes 4 thiol groups. The time course of the SH-titration is similar to that obtained when F-actin is depolymerized in a nucleotide free buffer. When actin is depolymerized in a buffer containing 10 equiv. of APPNP it also exposes 4 thiols. However, thiol-titration follows different kinetics. While one SH group reacts quickly the reaction of 3 others is retarded. We conclude that APPNP exhibits a shielding effect on part of the thiols for a period of time, while APPCP does not. In agreement with this, in the presence of APPNP yield of polymerization as well as stability against denaturation are distinctly higher than without added nucleotide or in the presence of APPCP. In line with this a hydrolysis product, most probably APPNH2, was associated with the filaments, as indicated by the replacement of tritiated ADP during polymerization, and from analysis of the attached nucleotide. Under the same conditions APPCP replaced tritiated ADP only to a small extent. The data indicate that APPNP interacts with monomeric actin much less than ATP and still less than ADP, but more so APPCP. APPNP is cleaved by actin ATPase and a hydrolysis product is incorporated into filaments.

19] Chemical derivatization of Ca2+-pump protein from skeletal muscle with N-substituted maleimides and 5-(2-iodoacetamidoethyl)aminonaphthalene 1-sulfonate

This chapter focuses on the chemical derivatizaton of Ca 2+ -pump protein from skeletal muscle with N-substituted Maleimides and 5-(2-Iodoacetamidoethyl)aminonaphthalene 1-Sulfonate. It has been difficult to achieve selective modification of a particular SH group, which is well-defined in terms of either its location or functional role, as a large number of SH groups are present in Ca 2+ ,Mg 2+ -ATPase of the sarcoplasmic reticulum (12–17 reactive SH groups in an ATPase polypeptide). In spite of the fact that the ATPase molecule has long been known to contain SH groups essential for the functioning of the transport system, and that their number has been suggested to be rather small, they have not yet been identified. SH groups were also utilized occasionally, as a target for covalent attachment of various probes to monitor conformational changes of transport ATPase during its catalytic cycle. The chapter describes simple procedures for selective derivatization of a limited number of specific SH groups with N-ethylmaleimide (MalNEt), and related fluorescent and paramagnetic probes, as well as a procedure for labeling with 5-(2-iodoacetamidoethyl)aminonaphthalene 1-sulfonate (IAEDANS), a fluorescent derivative of iodoacetamide.

Brdička-type catalytic currents of hemoglobins: Part III. pH dependence of the catalytic current - an indication of quaternary structure changes?

Changes of the catalytic current of bovine, ovine, rabbit, porcine, human, dog and pigeon hemoglobins (Hb), differing in the number of SH groups from 2 to 10 per tetramer, were studied in ammonia buffer solutions covering the pH range between 8.6 and 11.0. Two Hb concentrations were used: 5.0 × 10 −7 and 1.54 × 10 −6 M. In all cases, the pH rise was accompanied by an increase in the catalytic current in the form of two steps, until a maximum value of I catal at about pH 10.6–10.8 was attained, after which a decrease followed. A similar course of this relationship was observed for both concentrations of all the hemoglobins. The shape of the I catal = f(pH) curves resembled titration curves, from which common characteristic points were evaluated for all the hemoglobins, corresponding to pH values of about 9.8 ± 0.1 and 10.5 ± 0.1, done in a similar way as for p K. In the discussion, ionization of hemoglobins, complex formation with Co(II), denaturation, adsorption phenomena, in connection with the charge of Hb, quaternary structure changes, the accessibility and reactivity of SH groups and possible changes of the preparations prior to catalytic wave registration were taken into account. The most probable reason for formation of the second step in the I catal = f(pH) curves was considered to be the dissociation of deoxy-Hb tetramers to dimers. In the catalytic process, depending on the conditions at least two types of hemoglobin-Co(II) complex of different character may participate. The drop in I catal at high pH values was assumed to be the result of the competing action of ammonia on complexation processes involving hemoglobin and Co(II).

Studies on the reactivity of the essential sulfhydryl groups as a conformational probe for the fatty acid synthetase of chicken liver. Inactivation by 5,5'-dithiobis-(2-nitrobenzoic acid) and intersubunit cross-linking of the inactivated enzyme.

Fatty acid synthetase of chicken liver is rapidly and reversibly inactivated by 5,5'-dithiobis-(2-nitrobenzoic acid) (DTNB) at a rate (k2 = 132 mM-1 S-1 in 3 mM EDTA, 1% (v/v) glycerol, pH 7.0, at 25 degrees C) up to 2200 times higher than the reaction of this reagent with simple thiol compounds. The inactivation is caused by the reaction of the phosphopantetheine SH group, since it is protected competitively by either acetyl- or malonyl-CoA, and since the inactivated enzyme is unreactive with the phosphopantetheine label chloroacetyl-CoA but reactive with the cysteine reagent 1,3-dibromopropanone. Moreover, chloroacetyl-CoA prevents the modification of the rapidly reacting essential SH group by DTNB. The number of SH groups involved in inactivation was determined by correlating activity loss with the extent of reaction and by stopped-flow analysis of substrate (or chloroacetyl-CoA) protection. Values between 0.91 and 1.15 SH groups/dimer were obtained, indicating the presence of substoichiometric amounts of the prosthetic group in the fatty acid synthetase preparations used in this study. Inactivation of the synthetase by DTNB is strongly inhibited by increasing salt concentration and protected noncompetitively by NADP+ and NADPH. Treatment of the enzyme inactivated at low salt by salt, NADP+, or NADPH also effectively reduced cross-linking between enzyme subunits. The parallel effects of these treatments on the reaction with DTNB and subsequent dimerization are consistent with a minimum model of two discreet conformation states for fatty acid synthetase. In the low salt conformer, the phosphopantetheine and cysteine SH groups are juxtaposed, and the DTNB reaction (k2 approximately 132 mM-1 S-1) and dimerization are both facilitated. Transition to the high salt conformer by the above treatments is accompanied by an approximately 20-fold reduction of reactivity with DTNB (k2 = 6.8 mM-1 S-1) and reduced dimerization, due to spatial separation of the SH groups. During palmitate synthesis, the enzyme may oscillate between these conformation states to permit the reaction of intermediates at different active sites. Results obtained by studies on the effect of pH on DTNB inactivation implicate a pK of 5.9-6.1 for the essential SH group independent of salt concentration. This value is 1.5-1.8 pH units lower than the pK of 7.6-7.7 for CoA and may explain the 23-fold increase of the rate constant from a value of 0.3 mM-1 S-1 for CoA to that of the high salt conformer.

Evidence for the formation of a linear [3Fe-4S] cluster in partially unfolded aconitase.

Beef heart aconitase, as isolated under aerobic conditions, is inactive and contains a [3Fe-4S]1+ cluster. On incubation at pH greater than 9.5 (or treatment with 4-8 M urea) the color of the protein changes from brown to purple. This purple form is stable and can be converted back in good yield to the active [4Fe-4S]2+ form by reduction in the presence of iron. Active aconitase is converted to the purple form at alkaline pH only after oxidative inactivation. The Fe/S2- ratio of purple aconitase is 0.8, indicating the presence of [3Fe-4S] clusters. The number of SH groups readily reacting with 5,5'-dithiobis(2-nitrobenzoic acid) is increased from approximately 1 in the enzyme as isolated to 7-8 in the purple form, indicating a partial unfolding of the protein. On conversion of inactive aconitase to the purple form, the EPR signal at g = 2.01 (S = 1/2) is replaced by signals at g = 4.3 and 9.6 (S = 5/2). Mössbauer spectroscopy shows that purple aconitase has high-spin ferric ions, each residing in a tetrahedral environment of sulfur atoms. The three iron sites are exchange-coupled to yield a ground state with S = 5/2. Analysis of the data within a spin coupling model shows that J13 congruent to J23 and 2 J12 less than J13, where the Jik describe the antiferromagnetic (J greater than 0) exchange interactions among the three iron pairs. Comparison of our data with those reported for synthetic Fe-S clusters (Hagen, K. S., Watson, A. D., and Holm, R. H., (1983) J. Am. Chem. Soc. 105, 3905-3913) shows that purple aconitase contains a linear [3Fe-4S]1+ cluster, a structural isomer of the S = 1/2 cluster of inactive aconitase. Our studies also show that protein-bound [2Fe-2S] clusters can be generated under conditions where partial unfolding of the protein occurs.

Thiol-sensitive sites in cell adhesion. Decreased entry of SH-binding reagents into attached BHK cells.

Studies were carried out to learn more about the critical SH groups involved in cell spreading. Pretreatment of suspended baby hamster kidney (BHK) cells with 3 mM-iodoacetate or iodoacetamide for 10 min at 4 degrees C completely inhibited the ability of the cells to spread on fibronectin-coated substrata. If, however, BHK cells were permitted to attach and spread before being treated with the SH-binding reagents, and then harvested by trypsinization and assayed for spreading on fibronectin-coated substrata, there was no inhibition of cell spreading. The extent of prior attachment required before the cells became insensitive to the SH-binding reagents was tested and was found to occur early during the cell adhesion process, before any cell spreading was observed. In analytical experiments, there did not appear to be any difference in the total number of SH groups between suspended or spread cells as determined with 5,5'-dithiobis-(2-nitrobenzoic acid). The uptake of radiolabelled iodoacetate into intact spread cells, however, was found to be 3.5 times less than that found with suspended cells. On the other hand, the distribution of incorporated radioactivity into suspended and spread cells was similar. Most of the radioactivity (approximately 70%) was incorporated into small molecules (e.g. glutathione and cysteine), less (approximately 20%) was incorporated into cytoplasmic proteins, and the least incorporation (approximately 10%) was into the cell cytoskeleton. The data are interpreted to indicate there is a decreased permeability of spread cells to the SH-binding reagents.

Linear dependence of the polarographic catalytic current on the number of SH groups in the molecule for haemoglobins of different origin

Polarographic catalytic currents for solutions containing bovine, dog, human, rabbit, porcine or pigeon haemoglobins, and bovine or human methaemoglobins at several NH 3 and CoCl 2 concentrations were studied. At earlier elaborated conditions [9], for isomolar solutions of five haemoglobins of known amino-acids compositions, a linear correlation between the current of the first catalytic wave and the number of SH groups in the molecule was obtained, this being confirmed by a very high value of the correlation coefficient. No correlation could be stated using solutions of lower ammonia concentrations for the first and, under all conditions studied, for the second catalytic wave. On the basis of the linear dependence obtained, a number of SH groups in tetramers—and hence in the β-chain of dog haemoglobin and the α-chain of porcine haemoglobin—were found. The fact that the straight lines obtained do not intersect with the origin of coordinates is briefly discussed. Values of the standard deviation pointed to the use of low cobaltous chloride concentrations in the solution. It has also been stated that the magnitude of the catalytic current of haemoglobins depends on their methaemoglobin content, which should be connected with structural differences in these two species.

491—Linear dependence of polarographic catalytic current on the numeber of SH groups in the molecule for haemoglobins of different origin

Polarographic catalytic currents for solutions containing bovine, ovine, dog, human, rabbit, porcine or pigeon haemoglobins, and bovine or human methaemoglobins at several NH 3 and CoCl 2 concentrations were studied. At earlier elaborated conditions [9], for isomolar solutions of five haemoglobins of known amino-acid composition, a linear correlation between the current of the first catalytic wave and the number of SH groups in the molecule was obtained, this being confirmed by a very high value of the correlation coefficient. No correlation could be stated using solutions of lower ammonia concentrations for the first and, under all conditions studied, for the second catalytic wave. On the basis of the linear dependence obtained, a number of SH groups in tetramers—and hence in the ß-chain of dog haemoglobin and the α-chain of porcine haemoglobin—were found. The fact that the straight lines obtained do not intersect with the origin of coordinates is briefly discussed. Values of the standard deviation pointed to the use of low cobaltous chloride concentrations in the solution. It has also been stated that the magnitude of the catalytic current of haemoglobins depends on their methaemoglobin content, which should be connected with structural differences in these two species.

346 — Correlation between the SH group content of bovine or human haemoglobin and polarographic catalytic currents

Conditions were found under which the ratio of catalytic currents in the presence of human or bovine haemoglobin respectively amounted to nearly 3:1, corresponding to the ratio of the number of SH groups in their tetramers (or dimers). The data presented suggest that the composition of standard solutions used at present may be not the most suitable in searching for a direct correlation between the catalytic current and the content of SH groups of proteins. Advantages of the use of higher ammonia concentrations are briefly discussed.

346 — Correlation between the SH group content of bovine or human haemoglobin and polarographic catalytic currents

Conditions were found under which the ratio of catalytic currents in the presence of human or bovine haemoglobin respectively amounted to neary 3 : 1, corresponding to the ratio of the number of SH groups in their tetramers (or dimers). The data presented suggest that the composition of standard solutions used at present may be not the most suitable in searching for a direct correlation between the catalytic current and the content of SH groups of proteins. Advantages of the use of higher ammonia concentrations are briefly discussed.

Correlation between the SH group content of bovine or human haemoglobin and polarographic catalytic currents

Conditions were found under which the ratio of catalytic currents in the presence of human or bovine haemoglobin respectively amounted to nearly 3:1, corresponding to the ratio of the number of SH groups in their tetramers (or dimers). The data presented suggest that the composition of standard solutions used at present may be not the most suitable in searching for a direct correlation between the catalytic current and the content of SH groups of proteins. Advantages of the use of higher ammonia concentrations are briefly discussed.

Changes in the binding of copper in the plasma of molybdenum supplemented rats

After incubating plasma of Mo-supplemented rats (Mo-plasma) with 64Cu only part of it could be removed by dialysis against EDTA or histidine or by treatment with dithiocarbamate; this nondialyzable Cu was shown to be bound to albumin. The maximal amount of 64Cu bound this way equaled the Mo-induced increase in total plasma Cu. After addition of stable Cu dialysis of Mo-plasma against a histidine solution showed that no extra Cu became tightly bound, suggesting that the 64Cu binding was due to an exchange between added 64Cu and stable Cu already present. Incubating Mo-plasma with Hg compounds prevented 64Cu binding and released stable Cu, indicating that Cu in Mo-plasma was sulfhydryl bound. Part of the Mo in Mo-plasma was freely dialyzable. The remaining part was shown to be SH bound as well. The estimated atomic ratio of SH-bound Cu and Mo was unity. Molybdenum increased the number of SH groups in plasma, and for each Cu atom at least one SH group was calculated to be present.

Kinetics of thymidylate synthase inhibition by disulfides

The inactivation of thymidylate synthase (5,10-methylene-tetrahydrofolate: dUMP C- methyltransferase , EC 2.1.1.45) by a number of disulfides has been examined and found to be a second-order process. The apparent second-order rate constant was strongly influenced by the chemical structure of the disulfide. The data suggest that negatively charged functional groups decrease the reactivity of the disulfides and that positively charged groups enhance the reactivity. GSSG did not react with non-catalytic SH groups, since the number of SH groups of both GSSG-treated and untreated thymidylate synthase was the same. Several sulfhydryl compounds were tested for their ability to reactivate thymidylate synthase that had been inhibited by 2,2′-dithiodipyridine. Complete reactivation was obtained with either dithiothreitol or 2-mercaptoethanol. Reactivation by 2-mercaptoethanol was a second-order process.

Purification and Characterization of a 3,17 β-Hydroxysteroid Dehydrogenase from Streptomyces hydrogenans

3,17 β-Hydroxysteroid dehydrogenase has been enriched and purified from cytosol of Streptomyces hydrogenans. After ammonium sulfate precipitation and filtration on Sephadex G-100 the enzyme was finally purified by preparative gel electrophoresis and DEAE-Sephadex A-50 chro­matography. Polyacrylamide gel electrophoresis in the presence of sodium dodecylsulfate gave a single band of mobility corresponding to molecular weight of 70 200 ± 2 500. 3 β-. 17 β- as well as 20 β-hydroxy steroids were dehydrogenated by the enzyme in the presence of NAD+. The dehydrogenation proceeded faster than the reduction of the corresponding ketosteroids in the presence of NADH. The enzyme does not accent NADP+ or NADPH as co-substrates. The apparent Km values were calculated to be 11 μᴍ for 5 α-dihydrotestosterone, 20 μᴍ for testosterone ana 68 μᴍ for epiandrosterone in the NAD+-driven reaction, 1.8 x 10-4 m for NADH+ and 1.9 x 10-4 ᴍ for NADH. The catalytic activity was influenced by the ratio of NAD+/ATP. The inhibition by ATP appears to be of a competitive type with respect to NAD+ (Ki 1.15 x 10-3 ᴍ). After sucrose gradient centrifugation in a preparative ultracentrifuge the enzyme sediments with 4.1 ± 0.1 S as estimated in comparison to other proteins of known sedimentation coefficient. The isoelectric point was determined to be 3.9 with the LKB preparative isoelectric focusing col­umn (pH 2-11) and 4.1 with the analytical flat bed polyacrylamide isofocusing (pH 3 - 5). The number of SH groups was determined to be 2 mol/mol enzyme. In the presence of 6 M urea the fig­ure inceases to 3 mol SH/mol enzyme. In the presence of an excess of p-chloromercuribenzoate the enzyme activity decreases only partially.

Association between the Doubling Time of Various Cells and Tissues, and the SH-Content of Their Soluble Proteins

Abstract The num ber of SH-groups of the soluble proteins (Prot-SH) of 12 different types of animal tum ors and of cultivated chicken fibroblasts increases with decreasing doubling tim e (td). Between Prot-SH and td an inverse association was found with a significance level of 99.5%. After 30 min incubation in vitro, 4-hydroxypentenal (HPE) reacts with different am ounts of the Prot-SH of the investigated cells and tissues. It was found that the absolute, as well as the per­ cental amounts of HPE-reactive Prot-SH increase with decreasing td, each of the respective inverse associations was found to be highly significant. The tum ors were collected in three groups, each with a corresponding range of td and a mean value for Prot-SH . Taking the Prot-SH of the slowest growing tissue (DENA-hepatoma) as the reference value, the increments of Prot-SH were cal­ culated for each of the three groups and were found to be strikingly similar to the number of HPE-reactive Prot-SH. The single Prot-SH increm ents of the diverse cell and tissue types compared with DENA-hepatoma were found to be highly significantly and inversely associated with the respective Prot-SH decreases caused by HPE. Hence it may be concluded that HPE reacts preferably with those of the Prot-SH which are of some functional importance for tum or growth and, moreover that the attack on those thiols is the more effective the faster the tumor is growing.

Interaction of substrates and inhibitors with the homoserine dehydrogenase of kinase-inactivated aspartokinase I

The aspartokinase activity of the aspartokinase-homoserine dehydrogenase complex of Escherichia coli was affinity labeled with substrates ATP, aspartate, and feedback inhibitor threonine. Exchange-inert ternary adducts of Co(III)-aspartokinase and either ATP, aspartate or threonine were formed by oxidation of corresponding Co(II) ternary complexes with H2O2. The ternary enzyme-Co(III)-threonine adduct (I) had 3.8 threonine binding sites per tetramer, one-half that of the native enzyme. The binding of threonine to I was still cooperative as determined by equilibrium dialysis (nH = 2.2) or by studying inhibition of residual dehydrogenase activity (nH = 2.7). Threonine still protected the SH groups of I against 5,5'-dithiobis(2-nitrobenzoate) (DTNB) reaction but the number of SH groups reacting with thiol reagents (DTNB) was reduced by 1-2 per subunit in the absence of threonine. This suggests either that Co(III) is bound to the enzyme via sulfhydryl groups or that 1-2SH groups are buried or rendered inaccessible in I. The binding of threonine to sites not blocked by the affinity labeling produced changes in the circular dichroism of the complex comparable to changes produced by threonine binding to native enzyme and also protected against proteolytic digestion. The major conformational changes produced by threonine are thus ascribable to binding at this one class of regulatory sites. The interactions of kinase substrates with various aspartokinase-Co(III) complexes containing ATP, aspartate, or threonine and a threonine-insensitive homoserine dehydrogenase produced by mild proteolysis were studied. The inhibition of homoserine dehydrogenase by kinase substrates is not due to binding of these inhibitors at the kinase active site but was shown to be due to binding to sites within the dehydrogenase domain of the enzyme. L-alpha-Aminobutyrate, a presumed threonine analogue, also inhibits the dehydrogenase by binding at the same or similar sites in the dehydrogenase domain and not at threonine regulatory site.

DTNB modification of SH groups of isocitrate dehydrogenase from Bacillus stearothermophilus purified by affinity chromatography.

A new method of affinity chromatography using blue dextran-Sepharose 4B resin was established to purify NADP+-dependent isocitrate dehydrogenase [EC 1.1.1.42] from Bacillus stearothermophilus in high yield. The purified preparation was found to be homogeneous on disc gel electrophoresis. The SH groups of the enzyme were modified with 5,5'-dithiobis-(2-nitrobenzoic acid) (DTNB) to determine the number of SH groups per molecule and their contribution to the enzyme activity. One SH group was titrated with DTNB per subunit (the native enzyme consisted of two subunits) and after complete denaturation with 4 M guanidine-HCl the number of titratable SH groups remained unchanged. ORD and CD measurements showed that the alpha-helical conformation of the polypeptide backbone was unaffected by DTNB modification, though the near ultraviolet CD spectrum was evidently altered. The fluorescence derived from tryptophanyl residue(s) was quenched by the modification to 30% of the native level, which may indicate the presence of SH in the vicinity of tryptophanyl residue(s). A remarkable decrease of the enzyme activity was detected upon modification with DTNB, but there was some discrepancy between the rate of inactivation and that of modification of SH groups. The presence of substrate and Mg2+ gave partial protection against modification of the SH groups by DTNB. Complete protection of the native enzyme activity against heating at 65 degrees was observed in the presence of substrate and Mg2+, but the thermostability of the enzyme was markedly reduced by modification of the SH groups.

The modification of sulfhydryl groups of glutamine synthetase from Bacillus stearothermophilus with 5, 5'-dithiobis(2-nitrobenzoic acid).

The SH groups of glutamine synthetase [EC 6.3.1.2] from Bacillus stearothermophilus were modified with 5, 5'-dithiobis(2-nitrobenzoic acid) in order to determine the number of SH groups in the molecule as well as the effect of the modification on the enzyme activity. Three SH groups per subunit were detected after complete denaturation of the enzyme with 6 M urea, one of which was essential for the enzyme activity in view of its reactivity with 5, 5'-dithiobis(2-nitrobenzoic acid) on addition of MgCl2 with loss of the activity. The CD spectra of the modified enzyme in the near ultraviolet region changed from that of the native enzyme, indicating that aromatic amino acid residues were affected by modification of the SH group. The fluorescence derived from tryptophanyl residue(s) was quenched depending on the extent of modification of the SH group, suggesting that the tryptophanyl residue(s) was located in the proximity of the SH group. The thermostability of the enzyme was remarkably decreased by modification of the SH group.

Intramolecular crosslinking of tropomyosin via disulfide bond formation: evidence for chain register.

Rabbit skeletal muscle tropomyosin can be crosslinked in the native state by the use of 5,5'-dithiobis(2-nitrobenzoate), which forms disulfide bonds between the two subunits. Using polyacrylamide gel electrophoresis in sodium dodecyl sulfate we have shown that this crosslinking goes to completion over a wide range of protein concentration, ionic strength, and reagent concentration. Crosslinks are not formed in denaturing solvents such as sodium dodecyl sulfate and guanidine hydrochloride despite the fact that the same number of SH groups react as in the native state (2.3 +/- 0.2). The sedimentation coefficients of crosslinked and non-crosslinked samples are identical. Thus, crosslinks are formed between corresponding cysteines on different chains of the same molecule. This provides strong evidence for a model of chain interaction with both chains in register. Evidence has also been obtained that rabbit skeletal tropomyosin is composed only of alphaalpha and alphabeta subunits rather than a random mixture of chains.

Influence of L-thyroxine upon the ratio between protein-bound SH- and S-S-groups (author's transl)

In normal rats and in rats with mercury-induced tubular lesions without or with successive L-thyroxine treatment, the number of SH-groups and the total of SH- and S-S-groups in different segments of the nephron were calculated over a period of time by means of a photometric assay. The quantity of SH-groups was increased by low doses of sublimate but reduced by exogenous L-thyroxine, the whole of SH- and S-S-groups summing up to a constant quantity throughout. (Following application of high doses of sublimate, a loss of SH- as well as S-S-groups was observed, due to cellular destruction.) Connection and splitting of S-S-bridges, respectively, corresponded to changes in certain parameters of cell function, for instance resorptive capacity. A hypothesis is proposed stating that a fundamental effect of thyroid hormone is exerted by a biochemical action upon the ration between SH- and S-S-groups, permeability of the cellular and mitochondrial membranes being raised when the numerical proportion of S-S-bonds is high. Stimulation of enzymatic activity and metabolic processes would therefore result adaptively from the augmentation of substrate supply within the cell organells.