Inhibition Of ALAD Research Articles

STZ-induced diabetes in mice and heme pathway enzymes. Effect of allylisopropylacetamide and α-tocopherol

A frequent coexistence of diabetes and porphyria disease has been reported. Under normal conditions, porphyrin biosynthesis is well regulated to only form the amount of heme required for the synthesis of the various hemoproteins. The activity of some heme enzymes and rhodanese in streptozotocin (STZ) induced diabetic mice and in allylisopropylacetamide (AIA) induced experimental acute porphyria mice has been examined. The role of α-tocopherol (α-T), reported to prevent protein glycation in vitro, has also been investigated. AIA induced hepatic δ-aminolevulinic acid synthetase (ALA-S) activity in control animals but was ineffective in the diabetic group. α-Tocopherol did not modify ALA-S activity in either group. δ-Aminolevulinic acid dehydratase (ALA-D) and deaminase activities were significantly diminished both in liver and blood of diabetic animals. α-Tocopherol prevented inhibition of ALA-D, deaminase and blood rhodanese activities in diabetic animals but α-tocopherol by itself did not affect the basal levels of the enzymes studied. The potential use of a-tocopherol to prevent late complications of diabetes, including the onset of a porphyria like syndrome is considered.

Lead in hawks, falcons and owls downstream from a mining site on the Coeur d'Alene River, Idaho

Mining and smelting at Kellogg-Smelterville, Idaho, resulted in high concentrations of lead in Coeur d'Alene (CDA) River sediments and the floodplain downstream, where American Kestrels (Falco sparverius), Northern Harriers (Circus cyaneus), Red-tailed Hawks (Buteo jamaicensis), Great Horned Owls (Bubo virginianus), and Western Screech-owls (Otus kennicotti) nested. Nestling American Kestrels contained significantly higher (P=0.0012) blood lead concentrations along the CDA River (0.24 µg/g, wet wt) than the nearby reference area (0.087 µg/g). A 35% inhibition of blood δ-aminolevulinic acid dehydratase (ALAD) in nestling Northern Harriers (P=0.0001), 55% in nestling American Kestrels (P=0.0001) and 81% in adult American Kestrels (P=0.0004) provided additional evidence of lead exposure in the CDA River population. In nestling American Kestrels and Northern Harriers, ALAD activity was negatively correlated with lead in blood. An earlier report on Ospreys (Pandion haliaetus) showed slightly less inhibition of ALAD than in American Kestrels, but no significant reduction in hemoglobin or hematocrit and no negative influence on production rates. The adult and nestling American Kestrels along the CDA River contained about twice as much blood lead as Ospreys during the same years (adult 0.46 vs. 0.20 µg/g, and nestling 0.24 vs. 0.09 µg/g), but adults showed a 7.5% reduction in hemoglobin (P=0.0356) and nestlings an 8.2% reduction in hemoglobin (P=0.0353) and a 5.8% reduction in hematocrit (P=0.0482). We did not observe raptor deaths related to lead, and although the production rate for American Kestrels was slightly lower along the CDA River, we found no significant negative relation between productivity and lead. Limited data on the other raptors provide evidence of exposure to lead along the CDA River. Several traits of raptors apparently reduce their potential for accumulating critical levels of lead which is primarily stored in bones of prey species.

Delta-ALAD activity variations in red blood cells in response to lead accumulation in rock doves (Columba livia).

The enzyme {delta}-aminolevulinic acid dehydratase ({delta}-ALAD, E.C. 4.2.1.24), catalyses the second step of the haeme biosynthetic pathway and is required to maintain the haemoglobin and cytochrome content in red cells. {delta}-ALAD is not only found in bone marrow cells, the major site of haeme synthesis, but also in circulating erythrocytes and other tissues. An inverse correlation was found between {delta}-ALAD activity in red blood cells and lead concentration in the blood. The degree of {delta}-ALAD inhibition in erythrocytes has been widely accepted as a standard bioassay to detect acute and chronic lead exposure in humans and in avians. The value of this parameter as an indicator for environmental lead has been often reported in doves and Scanlon. In lead-treated rats, an increase in {delta}-ALAD activity in bone marrow cells and in blood samples was shown by radioimmunoassay at 5 and 9 days after the treatment. Similarly, the amount of {delta}-ALAD seems to be more sensitive to lead in avian species than in mammals, the usefulness of blood {delta}-ALAD activity as an index of lead exposure has already been questioned by Hutton in the pigeon and by Jaffe et al. in humans. The present investigation studied the toxic effects of leadmore » on rock dove red blood cell {delta}-ALAD activity in two situations: in doves treated with lead acetate in the laboratory and in doves exposed to the environment of Alcala de Henares. The final lead blood concentrations were lower in the environmental than in the laboratory doves. {delta}-ALAD activity in bone marrow cells and the relationships between lead accumulation and enzyme activity in red cells, are examined. 20 refs., 5 figs., 1 tab.« less

Assessment of the active-site requirements of 5-aminolevulinic acid dehydratase: evaluation of substrate and product analogs as competitive inhibitors

The enzyme 5-aminolaevulinic acid dehydrate (ALAD) is responsible for the synthesis of porphobilinogen (PBG) from two molecules of 5-aminolaevulinic acid (ALA). Porphobilinogen is an important committed intermediate in the biosynthesis of tetrapyrroles. The inhibition of ALAD from the purple bacterium Rhodopseudomonas sphaeroides was tested with various substrate and product analogues

Effects of water-borne copper and lead on the peripheral blood in the rosy barb,Barbus (Puntius) conchonius Hamilton

The Pb-induced abnormalities of hematopoiesis are primarily confined to the erythrocytes the leucocytes and platelets do not appear to be altered during chronic exposure. Pb effects in fishes show responses similar to those in mammals. The absence of erythrocyte {delta}-ALAD inhibition in fish exposed to Cd, Cu, Zn, and Hg indicated that this enzyme is quite specific for Pb. The objective of this work was to examine the effects of chronically sublethal concentrations of Cu and Pb on the peripheral blood parameters in the rosy barb, Barbus (Puntius) conchonius, a widely distributed freshwater bony fish.

Inhibition of liver, kidney, and erythrocyte δ-aminolevulinic acid dehydratase (porphobilinogen synthase) by gallium in the rat

Selective inhibition of enzymes in the heme biosynthesis pathway with concomitant urinary excretion of heme precursors serve as potentially important biological markers of chemical exposure and cell injury. Intratracheal administration of gallium arsenide particulate suspensions has been shown to result in inhibition of delta-aminolevulinic acid dehydratase (ALAD) in several tissues and increased excretion of the heme precursor aminolevulinic acid (ALA). This study was undertaken to evaluate in vivo the role of gallium alone in ALAD inhibition and increased urinary excretion of ALA. Male CD rats received a single ip injection of Ga2(SO4)3 at doses of 12.5, 25, 50, 100, and 200 mg Ga/kg. A dose-dependent inhibition of ALAD was observed 24 hr later in liver, kidney, and erythrocytes. After injection of 25 mg Ga/kg, maximal inhibition (42 to 49% of control) of ALAD occurred between 6 and 24 hr in liver and kidney with full recovery of activity at 96 hr. In erythrocytes, maximal inhibition (48% of control) occurred between 24 and 48 hr with recovery of activity at 96 hr. Mild to moderate renal proximal tubular necrosis in the pars recta was observed 24 hr after administration of 100 and 200 mg/kg, but no histopathologic changes were evident at lower doses. No consistent changes in urinary excretion of ALA were observed. Lineweaver-Burk analyses of renal and hepatic ALAD activities in the absence and presence of gallium indicated that the inhibition of ALAD by this element is noncompetitive (same Km, decreased Vmax). Gallium was shown to possess an inhibition constant (Ki) of approximately 3 microns for ALAD, similar to the Ki obtained for lead in other studies. Incubation of ALAD in vitro with gallium and lead, an active thiol group inhibitor, resulted in a greater inhibition of the enzyme. Further in vitro studies demonstrated the attenuation of gallium inhibition of hepatic and renal ALAD by zinc, suggesting that the mechanism of gallium action may involve competition for or displacement of zinc from the sulfhydryl group of the enzyme active site. Since ALAD inhibition occurred at doses at which no histopathologic changes were evident, the determination of ALAD activity in various tissues, including blood, may be of potential value as a biomarker of exposure/toxicity to metals such as gallium. The effect of chemical form and route of exposure of gallium and effects of other Group III metals on inhibition of ALAD and excretion of ALA is discussed.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of intratracheal gallium arsenide administration on δ-aminolevulinic acid dehydratase in rats: Relationship to urinary excretion of aminolevulinic acid

Exposure to gallium arsenide (GaAs) is a potential hazard in the semiconductor industry and there is a need for specific biological indicators of exposure/toxicity for this compound. These studies examined effects of GaAs exposure on the heme biosynthetic pathway enzyme δ-aminolevulinic acid dehydratase (ALAD). Male CD rats received GaAs suspensions at doses of 50, 100, or 200 mg/kg via a single intratracheal instillation. Six days after treatment a dose-dependent inhibition of blood ALAD was observed with activity decreasing to 5% of controls at the highest dose, with a concomitant marked increase in the urinary excretion of aminolevulinic acid (ALA). Inhibition of blood ALAD following administration of GaAs was maximal (30% of control) 3 to 6 days postexposure and returned to approximately control values on day 18. Urinary excretion of ALA was maximal 3 to 6 days postexposure and recovered toward control values at 18 days. Inhibition of kidney and liver ALAD following GaAs exposure was also evident. Intratracheal instillation of silica did not alter the activity of ALAD in blood, liver, or kidney. Marked increases in lung wet weight/body weight ratios were evident in lungs of silica- and GaAs-treated rats. Histopathological changes in the lungs were characterized by multifocal granuiomas following silica treatment and Type II pneumocyte hyperplasia following GaAs treatment; mild necrosis was evident in both groups. Rats treated with 100 mg/kg GaAs exhibited swelling of kidney proximal tubule mitochondria 6 days following exposure. Silica and GaAs exposure produced marked decreases in cumulative weight gain. The concentration of gallium required to achieve half-maximal inhibition of ALAD in vitro was 200-fold less in blood and 40-fold less in kidney and liver than that required for arsenite and the inhibition was partially prevented by excess zinc. These data suggest that gallium is the primary inhibitor of ALAD following dissolution of GaAs in vivo and that competition for or displacement of zinc from the enzyme active site may be involved in the mechanism of inhibition. The data also demonstrated the utility of including a particulate control group when assessing the chemical-induced toxicity of compounds administered intratracheally as particulate suspensions. Finally, measurement of heme precursors, e.g., ALA, in urine coupled with assay of red blood cell ALAD activity may be of value as an early biological indicator of GaAs exposure and/or toxicity.

Regulatory roles of high-affinity metal-binding proteins in mediating lead effects on delta-aminolevulinic acid dehydratase.

The present series of studies demonstrate that several high-affinity metal-binding proteins regulate essential metal availability and play a role in metal detoxication. Rat kidney PbBP and ZnMT were shown to mediate both the interaction of Zn and Pb to a Zn metalloenzyme, that is, ALAD. The mechanism for this interaction involves the donation of Zn from PbBP and ZnMT to ALAD and chelation of Pb to these proteins. In addition to activating ALAD via donation of Zn from ZnMT to the enzyme, kidney apothionein was shown to deactivate ALAD. Thus, these proteins may provide an intracellular reservoir for essential metals such as Zn and Cu, serving as a homeostatic control mechanism to readily dispense and sequester these cations to meet cellular metabolic requirements, such as the regulation of metalloenzymes, including ALAD. Although Pb does not effectively induce the synthesis of PbBP or ZnMT, these proteins, via binding of Pb, alter the biological activity of Pb toward a highly sensitive target molecule for Pb, that is, ALAD. Finally, when ALAD is utilized as a biological indicator for Pb exposure, the effect of ZnMT on Pb availability and ALAD activity must be taken into consideration. Tissue-specific differences in these high-affinity, metal-binding proteins and Zn status may alter the expected relationships between total tissue Pb concentrations and inhibition of ALAD.

Alterations in renal heme biosynthesis during metal nephrotoxicity.

The regulation of the heme biosynthetic pathway in the kidney by various metals has been reviewed. In addition, a study on the effects of lead on renal heme biosynthesis after acute treatment of rats has been reported. Chronic low-level lead exposure in rats results in relatively small effects on renal heme biosynthetic pathway enzymes. After acute treatment of rats with lead, no effects on ALAD or UROS and mild, transitory effects on ALAS and ferrochelatase are observed. The intracellular binding of lead within intranuclear inclusion bodies in the proximal tubule cells and to high-affinity cytosolic lead-binding proteins probably protects sensitive subcellular systems, such as the heme pathway, from lead toxicity. Chronic exposure to methyl mercury results in increased urinary excretion of uro- and coproporphyrins in rats, mediated via inhibition of ferrochelatase and UROS and stimulation of ALAS. A tissue-specific inhibition of ALAD occurs in the kidney after treatment of rats with indium. Acute treatment of rats with nickel, platinum, tin, antimony, bismuth, and cobalt results in induction of heme oxygenase, followed by decreased microsomal heme content and ALAS stimulation in the kidney.

Metal constitution of metallothionein influences inhibition of δ-aminolaevulinic acid dehydratase (porphobilinogen synthase) by lead

This study was undertaken to evaluate the effect of Zn and Cd pretreatment on the inhibition of delta-aminolaevulinic acid dehydratase (ALAD; porphobilinogen synthase, EC 4.2.1.24) by Pb. Male CD rats were pretreated with 200 mumol of Zn/kg s.c. (subcutaneously) or 18 mumol of Cd/kg s.c., 48 and 24 h before assay of ALAD. Pretreatment with Zn resulted in activation of hepatic and renal ALAD and attenuated the inhibition of this enzyme by Pb in vitro. Pretreatment with Cd increased hepatic ALAD activity, and the inhibitory effect of Pb on the hepatic enzyme was attenuated in this group. In contrast with the situation in liver, pretreatment with Cd did not affect the activity of renal ALAD and did not alter the inhibitory effect of Pb on the renal enzyme. The Pb IC50 (concentration causing half-maximal inhibition) values for hepatic and renal ALAD in Zn-pretreated rats and for hepatic ALAD in Cd-pretreated rats were increased above control, whereas the IC50 for renal ALAD in Cd-pretreated rats was unchanged. Cytosolic binding patterns for the three metals were assessed by gel-filtration chromatography and disclosed that 203Pb was co-eluted with Zn and Cd bound to liver and kidney Zn-thioneins and liver Cd,Zn-thionein, although minimal binding of 203Pb to kidney Cd,Zn-thionein was observed. Estimation of the molar ratio of metals bound revealed Cd/Zn ratios of 2 and 5 for Cd,Zn-thioneins from liver and kidney respectively. The inhibition of purified ALAD by Pb was also attenuated by addition of purified Zn-thioneins and Cd,Zn-thioneins from liver and kidney in the following order: liver Zn-thionein = kidney Zn-thionein greater than liver Cd,Zn-thionein much greater than kidney Cd,Zn-thionein. Thus liver and kidney Zn-thioneins and liver Cd,Zn-thionein with a low Cd/Zn ratio readily decrease the free pool of Pb available to interact with ALAD. These data also demonstrate that the capacity of metallothionein to alter the intracellular distribution of Pb and mediate the inhibition of ALAD by Pb is dependent on the tissue source and relative metal constitution of the metallothionein.

Kinetic parameters of the inhibition of red blood cell aminolevulinic acid dehydratase by triethyl lead and its reversal by dithiothreitol and zinc.

In chronic or acute exposure to triethyl lead, a de novo synthesis of aminolevulinic acid dehydratase (delta-ALAD) in bone marrow and an increased activity in circulating red blood cells can be demonstrated by activating the enzyme with dithiothreitol (DTT) and zinc. We determined the median inhibitory concentration and the apparent inhibition constant for triethyl lead on delta-ALAD. After dosing with triethyl lead, in vivo inhibition of ALAD only occurred at the high dose, but activation analysis in vitro showed increased ALAD activity to be present at all dose levels in a dose-dependent fashion. The use of an activation assay for red blood cell ALAD may have value as a bio-effects monitor of exposure to organic lead.

Kidney zinc-thionein regulation of δ-aminolevulinic acid dehydratase inhibition by lead

This study was undertaken to evaluate the ability of kidney Zn-thionein to regulate Zn availability to the Zn-dependent enzyme, δ-aminolevulinic acid dehydratase (ALAD), and mediate the effect of Pb on this enzyme. Male CD rats were pretreated with 200 μmol Zn/kg, sc, 48 and 24 h prior to assay of renal ALAD, which resulted in activation of renal ALAD and increased the resistance of this enzyme to inhibition by Pb in vitro. To determine the mechanism for this resistance, binding patterns of Zn and Pb in kidney cytosol were assessed. Rats were pretreated with Zn 48 and 24 h prior to injection of 203Pb (170 μCi/kg, ip). Kidneys were removed 4 h later and cytosol was fractionated on a Sephadex G-75 gel filtration column. Both 203Pb and Zn coeluted with the Zn-thionein fraction. Zn-thionein-I and -II, purified previously by DEAE anion-exchange chromatography, bound 203Pb in vitro. In another experiment, addition of purified Zn-thionein to reaction mixtures increased activity of purified bovine liver ALAD twofold and reversed inhibition of ALAD by Pb. Addition of apo-thionein to reaction mixtures partially prevented the inhibition of purified ALAD by Pb, indicating the biological significance of 203Pb chelation. Gel filtration of ALAD assay incubates containing 65Zn-thionein demonstrated that Zn is transferred from Zn-thionein to ALAD. Gel filtration of incubates containing 203Pb demonstrated that the presence of Zn-thionein alters the cytosolic binding pattern of Pb, with less bound to ALAD and more bound to Zn-thionein. The results demonstrate a dual function for Zn-thionein in mediating Pb inhibition of ALAD by a mechanism involving both donation of Zn to this Zn-requiring enzyme and chelation of Pb. These results also suggest that Zn-thionein may serve to regulate ALAD activity in vivo and mediate the inhibition of this enzyme by Pb.

Inhibition of δ-aminolevulinic acid dehydratase by trichloroethylene

Male Wistar rats (8 animals/group; 180–200 g) were exposed continuously to trichloroethylene (TRI) for 48 or 240 h or methylchloroform (1,1,1-trichloroethane: MC for 48 h at 50, 400 and 800 ppm. The inhibition of δ-aminolevulinic acid dehydratase (ALA-D) was examined in liver, blood and bone marrow of naive and phenobarbital pretreated animals exposed to TRI. A clear cut dose-effect relationship between the exposure concentration or duration of exposure and the inhibition of ALA-D activity was seen for rats exposed to TRI. In addition to this finding, significant interaction between TRI exposure and phenobarbital treatment was observed in the inhibition of ALA-D in liver and blood. MC did not produce inhibition. Trichloroacetic acid and trichloroethanol failed to inhibit the ALA-D activity in vitro. It seems that a metabolite(s) of TRI other than the above 2 substances may play a role in the inhibition of ALA-D. The inhibition of ALA-D (38% or 48% of the control in liver or in blood, respectively) observed after the 240 h exposure at 400 ppm to TRI was accompanied by the significant elevation of δ-aminolevulinic acid synthase (186% of the control) in liver and the increase in excretion of δ-aminolevulinic acid in urine (142% of the control). This occurred without an apparent weight loss, liver injury or hematological changes.

Postabsorptive Effect of Increased Dietary Zinc on Toxicity and Removal of Tissue Lead in Rats

Weanling male albino rats were fed a diet containing 12 ppm zinc and 200 ppm lead for 3 weeks. At the end of this time a representative number of samples were collected to determine tissue zinc and lead, inhibition of the lead-sensitive liver enzyme δ-aminolevulinic acid dehydratase (ALAD), and urinary δ-aminolevulinic acid (ALA). Dietary lead exposure was terminated, and the remaining rats were fed diets containing either 12 or 200 ppm zinc. Analyses were repeated at 5-day intervals over a 15-day period after lead exposure. As expected, inhibition of liver ALAD, excretion of urinary ALA and soft tissue lead content rapidly decreased after lead was removed from the diet approaching control levels by day 15. Although high dietary zinc increased the zinc content of plasma, liver and tibia, there was little or no therapeutic effect on recovery of liver ALAD, urinary ALA excretion or on the removal of lead from liver, kidney or tibia. Removal of red blood cell lead, however, was greater for rats fed the high zinc diet. Results of this study indicate that the post-absorptive interaction between zinc and lead is considerably less important than the previously reported intestinal interaction.

An Experimental Study to Protect the Inhibition of ALA-D due to Lead Exposure

An Experimental Study to Protect the Inhibition of ALA-D due to Lead Exposure

Use of Blood Lead and Erythrocyte θ-Amino Levulinic Acid Dehydratase Activity to Detect Lead Exposure of Fish Populations

The measurement of blood lead concentrations and inhibition of erythrocyte θ-amino levulinic acid dehydratase activity (ALA-D) has been used successfully to diagnose lead exposure in human populations. While blood lead is one of the best indicators of lead exposure, its measurement is expensive, time consuming, subject to bias through contamination and requires highly skilled personnel. The advantages of assaying ALA-D activity are those of cost, speed, sample size and simplicity. Since most organisms possess this enzyme in a variety of tissues, and since its activity is inhibited only by lead, there is potentially a large variety of aquatic species that may be used to monitor “biologically available” lead in aquatic ecosystems. Sessile and migratory species could integrate short-term fluctuations in waterborne lead and provide data on spatial and temporal variations. Fish are convenient organisms to sample and fish blood is a particularly rich source of ALA-D. Laboratory experiments have defined the optimum conditions for blood sampling and assaying ALA-D activity as well as the strong negative correlation between blood lead concentrations and ALA-D activity and between waterborne lead concentrations and ALA-D activity. Other toxic metals (e.g. Cu, Hg, Zn, Cd) and PCB's do not inhibit ALA-D, and factors that increase lead toxicity (e.g. decreased environmental pH) also increase lead uptake and the inhibition of ALA-D. Consequently, ALA-D activity provides a measure of both exposure and effect. Species variation in rates of lead uptake allows a selection of a suitable monitoring species for a given, situation. Preliminary surveys of Lake Ontario fish populations indicate that monitoring of ALA-D activity is technically simple and straightforward, the assay is much cheaper and faster than blood lead or whole body lead analyses, and activity is correlated to other measures of lead in fish.

Mechanisms of ALA-D inhibition by lead and of its restoration by zinc and dithiothreitol.

To induce the inhibition of ALA-D (delta-aminolaevulinic acid dehydratase) activity by lead in vitro, it is necessary to preincubate the enzyme fraction with lead ions and the Hb fraction (factors) together. The combination of two of the three (ALA-D fraction, lead acetate, and Hb fraction) in the preincubation has only a small effect on the activity. Lead preincubated with ALA-D and Hb fractions does not alter the affinity of the enzyme for the substrate, suggesting that the substrate can bind to the enzyme molecules (non-competitive inhibition). The restoration of activity by zinc with dithiothreitol is associated with the removal of lead from ALA-D fraction proteins to which it has bound in vivo and in vitro. The mode of the inhibitory action of tin on ALA-D is similar to that of lead because the inhibition is intensified by the addition of Hb fraction and is restored by heating. The inhibition concentration is, however, higher than that of lead. Of the three methods recovering decreased activity, heating is the most specific to detect inhibition by lead.

Relationships between distribution of lead in erythrocytes in vivo and in vitro and inhibition of ALA-D.

Proteins in the ALA-D (delta-aminolaevulinic acid dehydratase) fraction from gel filtration of erythrocyte supernatant (ES) have the highest affinity for lead among erythrocyte constituents in vivo and in vitro. It takes 20-40 hours for erythrocyte components to be equilibrated with lead added in vitro. AT low lead concentrations, under 60 micrograms/100 ml ES, the extent of decrease in ALA-D activity indicates the extent of lead saturation of ALA-D fraction proteins. The saturation is attained at 80-110 micrograms/100 ml ES. Although an appreciable amount of lead is also found in the haemoglobin fraction that contains certain factors concerned in ALA-D inhibition, lead responsible for inducing the inhibition is not bound to haemoglobin fraction proteins but to ALA-D fraction proteins. Of three treatments or agents recovering the enzyme from lead effects, zinc is the only one that can fully restore the inhibition.

Erythrocyte factors concerned in the inhibition of ALA-D by lead.

Erythrocyte factors are concerned in the inhibition of delta-aminolaevulinic acid dehydratase (ALA-D) by lead at 20 to 100 nM concentrations. The activity of the factors in detected in Hb fractions from Sephadex G-200 gel filtration of erythrocyte supernatant. After gel filtration of erythrocyte supernatant from a lead worker, 50% of lead is found in ALA-D fractions, although the fractions recover from ALA-D inhibition. The recovered activity is reinhibited if the enzyme fraction is preincubated with Hb fraction obtained from the same chromatography. Similarly obtained enzyme from a normal subject is also inhibited when it is preincubated with normal Hb fraction and lead acetate at 20 to 100 nM concentrations. The extent of the inhibition depends on the concentrations of Hb fraction and lead acetate preincubated. Reinhibition of lead worker enzyme with normal Hb fraction may be deleted not only by heating but also by zinc or DTT as well. Hb fraction heated at 60 degrees C for 5 min is also able to induce the lead-inhibition of the activity in ALA-D fraction. Half life of the factors is 26 min at 60 degrees C and 3 min at 80 degrees C.

Correlation of the physicochemical properties of metal ions with their activation and inhibition of human erythrocytic δ-aminolevulinic acid dehydratase (ALAD) in vitro

Using normal adult human whole blood hemolysates this study determined, in a dose-response fashion, the in vitro effects of Na +, Mg 2+, Al 3+, Mn 2+, Cu 2+, Ag +, Zn 2+, Cd 2+, Hg 2+, Ga 3+, In 3+, Sn 2+, Sn 4+, and Pb 2+ on normal erythrocytic ALAD. The effects of these 14 metal ions on erythrocytic ALAD 50% inhibited by Pb 2+ were also determined as was the ability of a maximum stimulatory concentration of Zn 2+ to prevent or reverse the effects of these metal ions on erythrocytic ALAD. The effects of these metal ions were then classified in terms of their oxidation state, characteristic coordination number, coordination geometry, and hard and soft Lewis acid characteristics in order to determine the physical and chemical properties associated with the ability of a metal ion to activate or inhibit erythrocytic ALAD and whether these properties are unique to a single metal ion. Preincubation studies established Zn 2+ to be the only metal ion to both activate erythrocytic ALAD and to prevent or reverse the Pb 2+-induced inhibition of erythrocytic ALAD in vitro even after prolonged contact with the enzyme. Therefore future investigations of the use of nontoxic salts of Zn 2+ as a prophylactic agent or therapeutic adjunct in the prevention or treatment of lead poisoning with its possibly toxic accumulation of ALA are proposed.