Biotransformation of terpenes from Stemodia maritima by Aspergillus niger ATCC 9142.

Incubation of stemodin (1) in cultures of Aspergillus niger ATCC 9142 resulted in the production of 2alpha,3beta,13-trihydroxystemodane (2), 2alpha,7beta,13-trihydroxystemodane (3) and 2alpha,13,16beta-trihydroxystemodane (4), while stemodinone (5) afforded 13,18-dihydroxystemodan-2-one (6) and 13,16beta-dihydroxystemodan-2-one (7). Four novel metabolites were obtained from the bioconversion of stemarin (8) by the fungus, namely 18-hydroxystemaran-19-oic acid (9), 7beta,18-dihydroxystemaran-19-oic acid (10), 7alpha,18,19-trihydroxystemarane (11) and 1beta-hydroxystemaran-19-oic acid (12). 19-N,N-Dimethylcarbamoxy-13-hydroxystemarane (13) was also transformed to afford 19-N,N-dimethylcarbamoxy-13,17xi,18-trihydroxystemarane (14).     

Stemodin-derived analogues with lipid peroxidation, cyclooxygenase enzymes and human tumour cell proliferation inhibitory activities

A series of analogues, derived from the antiviral and cytotoxic diterpene stemodin, were prepared and evaluated for their lipid peroxidation (LPO), cyclooxygenase enzyme-1 (COX-1) and -2 (COX-2), and tumour cell proliferation inhibitory activities. Oxidation of stemodin produced stemodinone, which was then converted to stemod-12-en-2-one. Reaction of the latter under Petrow conditions (bromine; silver acetate/pyridine) yielded mainly dibrominated abeo-stachanes. Solvolysis of the dibromo compounds gave products of hydrolysis, some with rearranged skeleta. In the lipid peroxidation inhibitory assay three of the compounds exhibited prominent activity. Interestingly, all the analogues showed higher COX-1 enzyme inhibition than COX-2. Although a few of the diterpenes limited the growth of some human tumour cell lines, most compounds induced proliferation of such cells.     

Isolation and structure of 13,18-dehydroexcelsin,a quassinoid,and glaucarubol from Ailanthus excelsa

A new quassinoid, 13,18-dehydroexcelsin and glaucarubol have been isolated from the bark of Ailanthus excelsa.     

Investigation of the importance of the C-2 oxygen function in the transformation of stemodin analogues by Rhizopus oryzae ATCC 11145

A new stemodinoside, stemodin-alpha-L-arabinofuranoside (5), was isolated from the plant Stemodia maritima. Incubation of stemodin (2) with Rhizopus oryzae ATCC 11145 gave 2 alpha,7 beta,13(S)-trihydroxystemodane (17) and 2 alpha,3 beta,13(S),16 alpha-tetrahydroxystemodane (18) whilst stemodinone (8) afforded 6 alpha,13(S)-dihydroxystemodan-2-one (19). The bioconversion of 2 beta,13(S)-dihydroxystemodane (10) by the fungus yielded 2 beta,7 beta,13(S)-trihydroxystemodane (20) whereas stemod-12-en-2-one (9) provided 7 beta,17-dihydroxystemod-12-en-2-one (21). The results provide useful information about the relationship between the functional groups of the substrates and their potential for bioconversion.     

Diterpenoids from Haplopappus rigidus

Fractionation of the aerial parts of Haplopappus rigidus Phil., directed by the brine shrimp lethality test (BST), has led to the isolation of two new diterpenoids, rigidusol and deacetylrigidusol. Their structures were established as 13-hydroxy-18-acetoxy-cis-cleroda-3,14-diene (8betaH, 10betaH, 19beta, 20alpha form) and 13,18-dihydroxy-cis-cleroda-3,14-diene (8betaH, 10betaH, 19beta, 20alpha form), respectively. Rigidusol exhibit moderate cytotoxic activity against human breast adenocarcinoma cell line MCF-7. Their structures were established by spectral data, in particular using 2D NMR spectroscopy (DEPT, DQF-COSY, HMQC and HMBC).     

Chromosomal mosaicism on amniotic interphase nuclei detected by multiprobe FISH

So far classical prenatal detection of chromosome aberrations has been limited to the evaluation of metaphase by means of time-consuming cytogenetic techniques. The MultiVision PGT test enables a simultaneous detection of aneuploidies of chromosomes 13,18, 21, X, and Y, even 24 h after amniocentesis. In the presented case, this test detected prenatally a chromosomal mosaicism 69,XYY[35]/46,XY[65]. This result was not confirmed after birth, by the same test on blood smear. The discrepancy is difficult to explain.     

Stemodane and stemarane diterpenoid hydroxylation by Mucor plumbeus and Whetzelinia sclerotiorum

Incubation of stemodin (1) with Mucor plumbeus ATCC 4740 resulted in the formation of 2alpha,6beta,13-trihydroxystemodane (2), 2alpha,3beta,13-trihydroxystemodane (3), 2alpha,11beta,13-trihydroxystemodane (4) and 2alpha,13,14-trihydroxystemodane (5), while stemodinone (7) afforded 6alpha,13-dihydroxystemodan-2-one (8) and 6alpha,12alpha,13-trihydroxystemodan-2-one (9). Metabolites obtained from the bioconversion of stemarin (11) were 8,13,19-trihydroxystemarane (12) and 2alpha,13,19-trihydroxystemarane (13). 19-N,N-Dimethylcarbamoxy-13-hydroxystemarane (14) was not transformed by the fungus. Stemodin (1) was incubated with Whetzelinia sclerotiorum ATCC 18687 to produce 2alpha,7beta,13-trihydroxystemodane (6) and 2alpha,11beta,13-trihydroxystemodane (4). Stemodinone (7) was converted to 7beta,13-dihydroxystemodan-2-one (10). Compounds 2, 4, 9, 10, 12 and 13 have not been previously reported.     

Bioconversion of Stemodia maritima diterpenes and derivatives by Cunninghamella echinulata var. elegans and Phanerochaete chrysosporium

Stemodane and stemarane diterpenes isolated from the plant Stemodia maritima and their dimethylcarbamate derivatives were fed to growing cultures of the fungi Cunninghamella echinulata var. elegans ATCC 8688a and Phanerochaete chrysosporium ATCC 24725. C. echinulata transformed stemodin (1) to its 7alpha-hydroxy- (2), 7beta-hydroxy- (3) and 3beta-hydroxy- (4) analogues. 2alpha-(N,N-Dimethylcarbamoxy)-13-hydroxystemodane (6) gave 2alpha-(N,N-dimethylcarbamoxy)-6alpha,13-dihydroxystemodane (7) and 2alpha-(N,N-dimethylcarbamoxy)-7alpha,13-dihydroxystemodane (8). Stemodinone (9) yielded 14-hydroxy-(10) and 7beta-hydroxy- (11) congeners along with 1, 2 and 3. Stemarin (13) was converted to the hitherto unreported 6alpha,13-dihydroxystemaran-19-oic acid (18). 19-(N,N-Dimethylcarbamoxy)-13-hydroxystemarane (14) yielded 13-hydroxystemaran-19-oic acid (17) along with the two metabolites: 19-(N,N-dimethylcarbamoxy)-2beta,13-dihydroxystemarane (15) and 19-(N,N-dimethylcarbamoxy)-2beta,8,13-trihydroxystemarane (16). P. chrysosporium converted 1 into 3, 4 and 2alpha,11beta,13-trihydroxystemodane (5). The dimethylcarbamate (6) was not transformed by this microorganism. Stemodinone (9) was hydroxylated at C-19 to give 12. Both stemarin (13) and its dimethylcarbamate (14) were recovered unchanged after incubation with Phanerochaete.     

From “naked” copper(II) ions and asymmetric macrocyclic complex linkers with a monodentate top and a bidentate top to molecular rectangle and infinite molecular rectangle strands

A molecular rectangle [Cu{CuL¹(NO₃)}(H₂O)(NO₃)]₂ (1) and two infinite molecular rectangle strands {[Cu{CuL¹(NO₃)}₂] · 2H₂O}_∞ (2) and [Cu{CuL²(ClO₄)}₂]_∞ (3) were prepared by reaction of “naked” Cu(II) ions with macrocyclic complex ligands CuL¹ for 1 and 2 and CuL² for 3 in metal-to-ligand molar ratios of 1:1, 1:2 and 1:2, respectively. L¹ and L² denote the dianions of diethyl 5,6,7,8,15,16-hexahydro-6,7-dioxodibenzo[1,4,8,11]tetraazacyclotetradecine-13,18-dicarboxylate and diethyl 5,6,7,8,15,16-hexahydro-15-methyl-6,7-dioxodibenzo[1,4,8,11]tetraazacyclotetradecine-13,18-dicarboxylate, respectively. The structures of 1-3 were determined by X-ray single-crystal analyses. CuL¹ in 1 and 2 and CuL² in 3 act as angular linkers with a monodentate coordination top and a bidentate one between two Cu(II) nodes to enclose the molecular rectangle of 1 and the rectangular subunits of 2 and 3. The angular shape, the monodentate top plus bidentate top coordination mode and the self-complementarity for π?π interactions of the macrocyclic complex linkers, the ratio between the reactants and the octahedral coordination geometry of the naked Cu(II) ions jointly determined the interesting structures of metallocyclophane 1 and 1D double chain coordination polymers 2 and 3. The cavities of the rectangular molecules of 1 are arranged into infinite strands so that parallel channels occur in the crystal. The molecules of 2 and 3 all pack parallel in the crystals.     

Secretoneurin facilitates endothelium-dependent relaxations in porcine coronary arteries

Secretoneurin enhances the adhesion and transendothelial migration properties of monocytes and is a part of the peptide family encoded by the secretogranin II gene. The expression of the secretogranin II gene is upregulated in senescent endothelium. The present study was designed to examine the effects of secretoneurin on endothelium-dependent responsiveness. Isometric tension was measured in rings (with or without endothelium) of porcine coronary arteries. Secretoneurin did not induce contraction of quiescent or contracted rings. In preparations contracted by U-46619, relaxation was observed with high concentrations of the peptide. This relaxation was endothelium dependent and reduced by the nitric oxide synthase inhibitor N(ω)-nitro-l-arginine methyl ester (l-NAME). It was abolished when the preparations were incubated with l-NAME in combination with the cyclooxygenase inhibitor indomethacin. The relaxation was not affected by the combination of 1-[(2-chlorophenyl)diphenylmethyl]-1H-pyrazole (TRAM-34) and 6,12,19,20,25,26-hexahydro-5,27:13,18:21,24-trietheno-11,7-etheno-7H-dibenzo[b,m][1,5,12,16]tetraazacyclotricosine-5,13-diiumditrifluoroacetate hydrate (UCL 1684), which abrogates endothelium-dependent hyperpolarizations. These results indicate that secretoneurin acutely induces relaxation through the activation of endothelial nitric oxide synthase (eNOS) and cyclooxygenase, with nitric oxide playing the dominant role. Prolonged (24 h) incubation with physiological concentrations of secretoneurin enhanced the relaxations to bradykinin and to the calcium ionophore A-23187, but this difference was not observed in preparations incubated with l-NAME or the calmodulin antagonist calmidazolium. Under these conditions, the relaxation to sodium nitroprusside remained unchanged. Incubation with secretoneurin significantly augmented the expression of eNOS and calmodulin as well as the dimerization of eNOS in cultures of porcine coronary arterial endothelial cells. These observations suggest that secretoneurin not only acutely causes but also, upon prolonged exposure, enhances endothelium-dependent relaxations.     

An energy budget for juvenile thick-lipped mullet, Crenimugil labrosus (Risso)

A series of experiments were carried out to construct an energy budget for juvenile thick lipped mullet, Crenimugil labrosus Risso. A partial factorial experimental design was used to examine the effects of temperature, fish size and meal size on growth. The maximum ration that the fish were able to ingest completely per day was found to be 0·8, 1·4 and 2·3% wet body weight (b.w.) at 13,18 and 23°C, respectively. Ingested maintenance requirements (M.R.) were estimated to be 137, 205 and 288 cal fish^-1 day^-1 at 13, 18 and 23°C, respectively. At 18deg; C, M.R. varied as 25 W^1.04 cal day^-1, where W= fish weight (g). Growth rate increased with increasing temperature. Maximal conversion efficiency was 21–24% and was achieved closer to the maximum ingested ration with increasing temperature. The relationship between respiration rate and W at 18deg; C for 3-20 g fish is described by: respiration rate (ml O₂ h^-1) = 0·128 W^0.976 The energy cost of apparent specific dynamic action at 18deg; C was found to vary between 5·1% and 23·6% of the calorific value of the ingested meal (1% wet b.w.) , mean (± S.E.)=10·2 ± 2·0%. Post mortem analyses of groups of fish fed 0·2, 0·8 and 1·5% wet b.w. meals showed a significant increase in total lipid and a significant decrease in water content with increasing ratio size. A negative correlation was found between body water content and total lipid (and calories). The mean assimilation efficiency (±s.e.) for 5–10 g mullet at 18deg; C was 73·9 ± 3·6%. The observations reported in this study were brought together to construct an energy budget for juvenile C. labrosus which was found to give a reliable prediction (within 10%) of energy demand and growth under the prevailing experimental conditions. Both gross (K₁) and net (K₂) growth efficiencies, based on energy values, increased with increasing ratio size up to satiation and were independent of temperature. The maximum values of K₁ and K₂ observed were 0·33 and 0·46, respectively. The third order efficiency (K₃) appeared to be independent of temperature and ration size; mean values ranged between 0·66 and 0·84.     

Levels of l-ascorbic acid and cadmium in the saphenous vein of patients with coronary artery disease are negatively correlated

BackgroundThe aim of this study was the simultaneous determination of levels of cadmium and l-ascorbic Acid (AA) in human saphenous vein (SV) used in coronary artery bypass grafting (CABG) and check whether there is a relationship between these levels.MethodsHuman SV were collected from 40 patients (20 men and 20 women; age, 40–75 years) at the time of routine coronary artery surgical revascularization. The concentration of cadmium in the tissue was determined according to the GF AAS—atomic absorption method. The concentration of AA was assayed in supernatant by FIA method with spectrophotometric detection.ResultsAA concentration (mean ± SD); men: 98,7 ± 13,18 μg/g tissue, women: 96,06 ± 11,98 μg/g tissue. Cadmium concentration(mean ± SD); men: 309 ± 103,71 ng/g tissue, women: 348,5 ± 255,71 ng/g tissue. Correlations among concentrations of AA and cadmium were insignificant negative in the group of men (Pearson r = −0,1504, p = 0,5269) and in the group women (Pearson r = −0339, p = 0144).ConclusionsNegative correlations among concentrations of AA and cadmium in human SV obtained in our study may indicate a protective effect of this vitamin in relation to toxic cadmium.     

The effect of divalent cations on bovine spermatozoal adenylate cyclase activity.

The effect of divalent cations on bovine sperm adenylate cyclase activity was studied. Mn2+, Co2+, Cd2+, Zn2+, Mg2+ and Ca2+ were found to satisfy the divalent cation requirement for catalysis of the bovine sperm adenylate cyclase. These divalent cations in excess of the amount necessary for the formation of the metal-ATP substrate complex were found to stimulate the enzyme activity to various degrees. The magnitude of stimulation at saturating concentrations of the divalent cations was strikingly greater with M2+ than with either Ca2+, Mg2+, Zn2+, Cd2+ or Co2+. The apparent Km was lowest for Zm2+ (0.1 - 0.2 mM) than for any of the other divalent cations tested (1.2 - 2.3 mM). The enzyme stimulation by Mn2+ was decreased by the simultaneous addition of Co2+, Cd2+, Ni2+ and particularly Zn2+ and Cu2+. The antagonism between Mn2+ and Cu2+ or Zn2+ appeared to have both competitive and non-competitive features. The inhibitory effect of Cu2+ on Mn2+-stimulated adenylate cyclase activity was prevented by 2,3-dimercaptopropanol, but not by dithiothreitol, L-ergothioneine, EDTA, EGTA or D-penicillamine. Ca2+ at concentrations of 1-5 mM was found to act synergistically with Mg2+, Zn2+, Co2+ and Mn2+ in stimulating sperm adenylate cyclase activity. The Ca2+ augmentation of the stimulatory effect of Zn2+, Co2+, Mg2+ and Mn2+ appeared to be specific.     

Characterization and crystal structure of cadmium(II) halide complexes with amino acids and their derivatives VI. The comparison of crystal structures of cadmium(II) halide complexes with three kinds of piperidine carboxylic acids

Six cadmium(II) halide complexes with dl-piperidine-2-carboxylic acid (DL-Hpipe-2), dl-piperidine-3-carboxylic acid (DL-Hpipe-3), and piperidine-4-carboxylic acid (Hpipe-4), have been prepared and characterized by means of IR and Raman spectra and thermal analysis. The crystal structures of [CdCl2(DL-Hpipe-2)(H2O)], [CdBr2(DL-Hpipe-3)], and [CdCl2(Hpipe-4)] have been determined by X-ray diffraction. These three complexes have one-dimensional polymer structures bridged by halide atoms. The crystal of [CdCl2(DL-Hpipe-2)(H2O)] is orthorhombic with the space group Pca2(1). The cadmium atom is in an octahedral geometry, ligated by a carboxyl oxygen atom, two bridging chlorine atoms, a terminal chlorine atom, a water molecule and a carboxyl oxygen atom of a neighboring molecule. The carboxyl oxygen atoms of DL-Hpipe-2 are coordinated to two cadmium atoms. The unit cell consists of two types of one-dimensional polymer structures: [CdCl2(D-Hpipe-2)(H2O)] and [CdCl2(L-Hpipe-2)(H2O)]. Therefore, it is better to write [CdCl2(DL-Hpipe-2)(H2O)] as [CdCl2(D-Hpipe-2)(H2O)][CdCl2(L-Hpipe-2)(H2O)]. The crystal structure of [CdBr2(DL-Hpipe-3)] is monoclinic with space group P2(1). The cadmium atom is in a distorted octahedral geometry ligated by two carboxyl oxygen atoms and four bridging bromine atoms. This complex consists of either D-Hpipe-3 or L-Hpipe-3. Therefore [CdBr2(DL-Hpipe-3)] is written as [CdBr2(D or L-Hpipe-3)]. The crystal of [CdCl2(Hpipe-4)] is monoclinic with space group P2(1)/n. The structure is similar to that of [CdBr2(D or L-Hpipe-3)].     

酵母RNA聚合酶ⅡRpb2和Rpb3两亚基间相互作用位点的定位

为研究S .pombeRNApolⅡ各亚基间体内装配成复合体的机制 ,本文首次用酵母双杂交系统鉴定了Rpb2和Rpb3两亚基间体内相互作用的位点。首先将Rpb2的 4个片段克隆至Gal4BD表达载体pAS2上 ,构建BD Rpb2片段融合蛋白重组质粒 ;同时将Rpb3克隆至Gal4AD表达载体pGADGH上 ,构建AD Rpb3融合蛋白重组质粒。其次 ,将pGADGHRpb3分别与pAS2Rpb2各片段重组质粒共转化到受体酵母菌Y1 90感受态细胞内 ,筛选并鉴定β gal活性阳性 (β gal+)的共转化子。最后 ,将β gal+共转化子中的Rpb2片段进行序列分析并进行同源序列比较确定其在Rpb2中的位置。结果表明 ,Rpb2与Rpb3相互作用的位点位于Rpb2的 90 2～ 989aa肽段内     

Interrelationship between oxygen consumption,superoxide anion and hydrogen peroxide formation in phagocytosing guinea pig polymorphonuclear leucocytes

The paper presents an experimental procedure for a simultaneous assay of oxygen consumption, O2- release and H2O2 accumulation at a very early stage of the respiratory burst that is induced by phagocytosis in guinea pig polymorphonuclear leucocytes. The main findings are as follows: (a) The oxygen consumption that is measurable does not correspond to all oxygen that is reduced. The relationship between the actual oxygen consumed and the amount that is reduced depends on the fate of the intermediate products O2- and H2O2. (b) O2- is measurable extracellularly by the reduction of cytochrome c. When cytochrome c oxidizes the extracellular O2-, molecular oxygen is formed. This fact is shown by a decrease of oxygen consumption. The molar ratio between the O2- detected and the oxygen given back is 1. (c) The amount of O2- released from the cells accounts for only a small part of oxygen actually reduced. (d) H2O2 is detectable only in the presence of NaN3. In this condition almost all oxygen consumed is recovered in the form of H2O2. The molar ratio O2/H2O2 is near unity. The amount of H2O2 derived from dismutation of O2- released is only an aliquot of the total H2O2 accumulated. Thus, most of H2O2 is derived from intracellular sources. (e) In the absence of inhibitors of H2O2 degrading reactions, no detectable accumulation of peroxide occurs. Under these conditions, the main part of H2O2 formed is degraded in almost equal amount by catalase and myeloperoxidase, while only a small aliquot is degraded by NaN3 insensitive reactions.     

A chemoenzymatic route to mannosamine derivatives bearing different N-acyl groups

The chemoenzymatic route to 2-deoxy-2-propionamido-D-mannose (1b), 2-butyramido-2-deoxy-D-mannose (2b) and 2-deoxy-2-phenylacetamido-D-mannose (3b) involved N-acylation of 2-amino-2-deoxy-D-glucose followed by alkaline C-2 epimerization and selective microbial removal of the epimers with gluco-configuration. The latter step employed whole cells of Rhodococcus equi A4 able to degrade 2-deoxy-2-propionamido-D-glucose (1a), 2-butyramido-2-deoxy-D-glucose (2a) and 2-deoxy-2-phenylacetamido-D-glucose (3a) but inactive towards the corresponding manno-isomers. The metabolism of the gluco-isomers probably involved phosphorylation and subsequent deacylation. 2-Acetamido-2-deoxy-6-O-phospho-D-glucose amidohydrolase [EC 3.5.1.25] but not 2-acetamido-2-deoxy-D-glucose amidohydrolase was detected in the cell extract, the former enzyme being partially purified (15.8-fold with an overall yield of 18.1% and a specific activity of 0.95 units mg-1 protein). According to SDS-PAGE electrophoresis, gel filtration and mass spectrometry, the enzyme was a monomer with an apparent molecular mass of approximately 42 kDa. The optimum temperature and pH of the enzyme were 60 degrees C and 8.0-9.0, respectively. 2-Acetamido-2-deoxy-6-O-phospho-D-glucose and 2-acetamido-2-deoxy-6-O-sulfo-D-glucose but not 2-acetamido-2-deoxy-1-O-phospho-D-glucose or 2-acetamido-2-deoxy-D-glucose were substrates of the enzyme. Its activity was slightly inhibited by the addition of 1 mM Al3+, Ca2+, Co2+, Cu2+, Mn2+ or Zn2+ and activated by 1 mM Mg2+. The concentrated enzyme is highly stable at 4 degrees C in the presence of 0.1 M ammonium sulfate.     

Metabolic response of Platynota stultana pupae to controlled atmospheres and its relation to insect mortality response

The metabolic responses of Platynota stultana pupae to reduced O(2), elevated CO(2), and their combinations were investigated using microcalorimetry, and mortality of pupae under elevated CO(2) atmospheres was correlated with metabolic responses. The metabolic heat rate decreased slightly with decreasing O(2) concentration until a critical O(2) concentration (P(c)) below which the heat rate decreased rapidly. The P(c) increased with temperature. The percentage decreases of metabolic heat rate were comparable to the percentage decreases of O(2) consumption rate (RO(2)) at 10, 8, 6, and 4% O(2), but were smaller at 2 and 1% O(2). The metabolic heat rate decreased rapidly at 20% CO(2) relative to 0% CO(2), with little to no further decrease between 20 and 79% CO(2). The percentage decreases of RO(2) under 20 and 79% CO(2) at 20 degrees C were comparable to the percentage decreases of metabolic heat rates. The additive effects of subatmospheric O(2) and elevated CO(2) levels on reducing metabolic heat rate were generally fully realized at combinations of /=4% O(2), but became increasingly overlapped as the O(2) concentration decreased and the CO(2) concentration increased. The high susceptibility of pupae to elevated CO(2) at high temperature was correlated with high metabolic heat rate. The metabolic responses of pupae to reduced O(2) concentrations included metabolic arrest and anaerobic metabolism. The net effect of elevated CO(2) on the pupal respiratory metabolism was similar to that of reduced O(2); however, mechanisms other than the decrease of metabolism were also contributing to the toxicity of CO(2).     

The effects of Mg2+ on rat liver microsomal Ca2+ sequestration

The effects of Mg2+ on the hepatic microsomal Ca2(+)-sequestering system was tested. Ca2(+)-ATPase activity and Ca2+ uptake were both dependent on the concentration of free Mg2+, reaching maximum levels at 2 mM. The effects of Mg-ATP were also influenced by the concentration of free Mg2+, being maximally effective at a ratio of 1:1. The results suggest that Mg2+ influences Ca2+ sequestration at various steps, namely in addition to forming the substrate of the Ca2(+)-ATPase reaction, Mg-ATP, Mg2+ stimulates the reaction at an additional step, as indicated by its stimulatory effect on the Ca2(+)-ATPase reaction and on Ca2+ uptake, even at optimal Mg-ATP levels. The stimulatory effect of Mg2+ was evident at various pH levels tested, and it was nucleotide specific. The stimulatory effect of Mg2+ might be exerted at the dephosphorylation step of the enzymatic reaction or at an other, yet undefined, site. The results demonstrate a plural effect of Mg2+ on the hepatic microsomal sequestration system. This indicates that, depending on its magnitude, changes in Mg2+ distribution might influence cytosolic Ca2+ levels.