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Karl T Kelsey - One of the best experts on this subject based on the ideXlab platform.

  • lead burden and psychiatric symptoms and the modifying influence of the δ aminolevulinic acid dehydratase ALAD polymorphism the va normative aging study
    American Journal of Epidemiology, 2007
    Co-Authors: Karl T Kelsey, P Rajan, Joel Schwartz, David C Bellinger, Jennifer Weuve, David Sparrow, Avron Spiro
    Abstract:

    Alterations in mood among the elderly not only disrupt their regular physical, mental, and social functioning but also may place them at a higher risk of developing clinically relevant mental disorders (1, 2) and cardiovascular disease (3–5). In particular, depressive symptoms are associated with a higher risk of functional decline (6, 7) and self-neglect (8). Lead exposure is associated with increased psychiatric symptomatology in some occupationally exposed adults (9–11). However, the results of other occupational studies are inconsistent with these findings and report that cumulative and concurrent lead burden is not associated with psychiatric symptoms (12, 13). Few studies have addressed the impact of low-level cumulative and concurrent lead burden on psychiatric symptoms among the general population. In our evaluation of lead and psychiatric symptoms in environmentally exposed older men, higher chronic and cumulative lead burden (measured by lead levels in the patella and tibia) was significantly associated with phobic anxiety symptoms and a composite index of depression, anxiety, and phobic anxiety symptoms (14). In the same study, higher levels of concurrent lead exposure (measured by lead levels in blood) were also significantly associated with the composite symptom index, suggesting that even modest levels of lead exposure may increase the risk of developing psychiatric symptoms. For this study, we expanded upon our earlier work by incorporating newly available data from up to two additional assessments of both psychiatric symptoms and lead burden. In addition, we evaluated the influence of a polymorphism in the gene encoding δ-aminolevulinic acid dehydratase (ALAD) on the relation of lead to mood. This ALAD polymorphism (rs1800435) encodes three distinctively charged erythrocyte isoenzymes in the heme synthesis pathway: ALAD 1-1 (52.1 mIU/g hemoglobin), ALAD 1-2 (49 mIU/g hemoglobin), and ALAD 2-2 (54.6 mIU/g hemoglobin) (15). In comparison with ALAD 1-1 carriers, ALAD 1-2/2-2 carriers have been reported to have a higher percentage of lead bound to the ALAD protein (16), potentially altering the kinetic distribution of lead to target organs (17, 18). However, ALAD genotype modulation of lead toxicokinetics is poorly understood, and prior studies on the relation between ALAD genotype, lead, and neurologic outcomes have reported inconsistent findings (19–21). We anticipated that the dose-response association between biomarkers of lead burden and psychiatric symptoms would be steeper among men with the wild-type genotype (ALAD 1-1) in comparison with men with a variant allele (ALAD 1-2/2-2), reflecting a potential increase in the retention of lead in blood among this latter group and therefore decreased bioavailability for crossing the blood-brain barrier.

  • A delta-aminolevulinic acid dehydratase (ALAD) polymorphism may modify the relationship of low-level lead exposure to uricemia and renal function: the normative aging study.
    Environmental health perspectives, 2003
    Co-Authors: Karl T Kelsey, Joel Schwartz, David Sparrow, Scott T. Weiss
    Abstract:

    In this study we investigated whether a known delta-aminolevulinic acid dehydratase (ALAD) exon 4 polymorphism has a modifying effect on the association of blood or bone lead level with uricemia and indices of renal function among middle-aged and elderly men. We performed a cross-sectional study of subjects who participated between 1991 and 1995 in the Department of Veterans Affairs Normative Aging Study. Information on blood lead levels, bone lead levels (measured by K-shell X-ray fluorescence), serum uric acid, serum creatinine, estimated creatinine clearance, and ALAD polymorphism status was available in 709 subjects. Regression models were constructed to examine the relationships of serum uric acid, serum creatinine, and estimated creatinine clearance to blood or bone lead level, stratified by genotype. We also adjusted for age, body mass index, blood pressure, smoking, alcohol consumption, and ingestion of analgesic medications (n = 638). Of the 709 subjects, 7 (1%) and 107 (15%) were homozygous and heterozygous for the variant (ALAD-2) allele, respectively. The mean (range) serum uric acid and creatinine levels were 6.5 (2.9-10.6) and 1.2 (0.6-2.5) mg/dL. No significant differences were found in serum uric acid, serum creatinine, or estimated creatinine clearance by ALAD genotype. However, after adjusting for other potential confounders, we found a significant linear relationship between serum uric acid and patella bone lead (p = 0.040) among the ALAD 1-2/2-2 genotype individuals above a threshold patellar lead level of 15 micro g/g. In contrast, among the wild-type (ALAD 1-1) individuals, there was a suggestion of a significant linear relationship of serum uric acid with patella bone lead (p = 0.141), but only after a threshold of 101 micro g/g. There was evidence of a significant (p = 0.025) interaction of tibia lead with genotype (ALAD 1-1 vs. ALAD 1-2/2-2) regarding serum creatinine as an outcome, but in the same linear regression model tibia lead alone was not a significant predictor of serum creatinine. Conversely, for estimated creatinine clearance, patella lead, but not the interaction of patella lead with genotype, was a significantly independent predictor (p = 0.026). Our findings suggest that ALAD genotype may modify the effect of lead on the renal excretion of uric acid as well as overall renal function among middle-aged and elderly men who had community (nonoccupational) exposures to lead. Additional research is needed to ascertain whether this constitutes a true gene-environment interaction and, if so, its clinical impact.

  • The delta-aminolevulinic acid dehydratase (ALAD) polymorphism and bone and blood lead levels in community-exposed men: the Normative Aging Study.
    Environmental health perspectives, 2001
    Co-Authors: Yawen Cheng, David Sparrow, Scott T. Weiss, Karl T Kelsey
    Abstract:

    Recent research has indicated that a polymorphic variant of delta-aminolevulinic acid dehydratase (ALAD) may influence an individual's level of lead in bone and blood and, as a result, may also influence an individual's susceptibility to lead toxicity. In this study, we investigated whether this ALAD polymorphism is associated with altered levels of lead in bone and blood among 726 middle-aged and elderly men who had community (nonoccupational) exposures to lead. We measured levels of blood and bone lead by graphite furnace atomic absorption spectroscopy and a K X-ray fluorescence (KXRF) instrument, respectively. We determined the ALAD MspI polymorphism in exon 4 by a polymerase chain reaction restriction fragment length polymorphism (RFLP). Of the 726 subjects, 7 (1%) and 111 (15%) were, respectively, homozygous and heterozygous for the variant allele. The mean (SD) of blood lead (micrograms per deciliter), cortical bone (tibia) lead (micrograms per gram), and trabecular bone (patella) lead (micrograms per gram) were 6.2 (4.1), 22.1 (13.5), and 31.9 (19.5) in subjects who did not have the variant allele (ALAD 1-1), and 5.7 (4.2), 21.2 (10.9), and 30.4 (17.2) in the combined subjects who were either heterozygous or homozygous for the variant allele (ALAD 1-2 and ALAD 2-2). In multivariate linear regression models that controlled for age, education, smoking, alcohol ingestion, and vitamin D intake, the ALAD 1-1 genotype was associated with cortical bone lead levels that were 2.55 microg/g [95% confidence interval (CI) 0.05-5.05] higher than those of the variant allele carriers. We found no significant differences by genotype with respect to lead levels in trabecular bone or blood. In stratified analyses and a multivariate regression model that tested for interaction, the relationship of trabecular bone lead to blood lead appeared to be significantly modified by ALAD genotype, with variant allele carriers having higher blood lead levels, but only when trabecular bone lead levels exceeded 60 microg/g. These results suggest that the variant ALAD-2 allele modifies lead kinetics possibly by decreasing lead uptake into cortical bone and increasing the mobilization of lead from trabecular bone.

  • interaction of blood lead and delta aminolevulinic acid dehydratase genotype on markers of heme synthesis and sperm production in lead smelter workers
    Environmental Health Perspectives, 1998
    Co-Authors: Bruce H Alexander, Karl T Kelsey, Harvey Checkoway, Paola Costamallen, Elaine M Faustman, James S Woods, C Van Netten, Lucio G Costa
    Abstract:

    The gene that encodes gamma-aminolevulinic acid dehydratase (ALAD) has a polymorphism that may modify lead toxicokinetics and ultimately influence individual susceptibility to lead poisoning. To evaluate the effect of the ALAD polymorphism on lead-mediated outcomes, a cross-sectional study of male employees from a lead-zinc smelter compared associations between blood lead concentration and markers of heme synthesis and semen quality with respect to ALAD genotype. Male employees were recruited via postal questionnaire to donate blood and urine for analysis of blood lead, zinc protoporphyrin (ZPP), urinary coproporphyrin (CPU), and ALAD genotype, and semen samples for semen analysis. Of the 134 workers who had ALAD genotypes completed, 114 (85%) were ALAD1-1 (ALAD1) and 20 (15%) were ALAD1-2 (ALAD2). The mean blood lead concentrations for ALAD1 and ALAD2 were 23.1 and 28.4 microg/dl (p = 0.08), respectively. ZPP/heme ratios were higher in ALAD1 workers (68.6 vs. 57.8 micromol/ml; p = 0.14), and the slope of the blood lead ZPP linear relationship was greater for ALAD1 (2.83 vs. 1.50, p = 0.06). No linear relationship between CPU and blood lead concentration was observed for either ALAD1 or ALAD2. The associations of blood lead concentration with ZPP, CPU, sperm count, and sperm concentration were more evident in workers with the ALAD1 genotype and blood lead concentrations >/= 40 microg/dl. The ALAD genetic polymorphism appears to modify the association between blood lead concentration and ZPP. However, consistent modification of effects were not found for CPU, sperm count, or sperm concentration.

  • delta aminolevulinic acid dehydratase genotype modifies four hour urinary lead excretion after oral administration of dimercaptosuccinic acid
    Occupational and Environmental Medicine, 1997
    Co-Authors: Brian S Schwartz, Walter F Stewart, Pornchai Sithisarankul, Paul Timothy Strickland, Karl T Kelsey
    Abstract:

    OBJECTIVES: Previous research suggests that binding of lead by delta-aminolevulinic acid dehydratase (ALAD) may vary by ALAD genotype. This hypothesis was tested by examining whether ALAD genotype modifies urinary lead excretion (DMSA chelatable lead) after oral administration of dimercaptosuccinic acid (DMSA). METHODS: 57 South Korean lead battery manufacturing workers were given 5 mg/kg oral DMSA and urine was collected for four hours. Male workers were randomly selected from two ALAD genotype strata (ALAD1-1, ALAD1-2) from among all current workers in the two plants (n = 290). Subjects with ALAD1-1 (n = 38) were frequency matched with subjects with ALAD1-2 (n = 19) on duration of employment in the lead industry. Blood lead, zinc protoporphyrin, and plasma aminolevulinic acid concentrations, as well as ALAD genotype, duration of exposure, current tobacco use, and weight were examined as predictors or effect modifiers of levels of DMSA chelatable lead. RESULTS: Blood lead concentrations ranged from 11 to 53 micrograms/dl, with a mean (SD) of 25.4 (10.2) micrograms/dl. After 5 mg/kg DMSA orally, the workers excreted a mean (SD) 85.4 (45.0) micrograms lead during a four hour urine collection (range 16.5-184.1 micrograms). After controlling for blood lead concentrations, duration of exposure, current tobacco use, and body weight, subjects with ALAD1-2 excreted, on average, 24 micrograms less lead during the four hour urine collection than did subjects with ALAD1-1 (P = 0.05). ALAD genotype seemed to modify the relation between plasma delta-aminolevulinic acid (ALA) and DMSA chelatable lead. Workers with ALAD1-2 excreted more lead, after being given DMSA, with increasing plasma ALA than did workers with ALAD1-1 (P value for interaction = 0.01). CONCLUSIONS: DMSA chelatable lead may partly reflect the stores of bioavailable lead, and the current data indicate that subjects with ALAD1-2 have lower stores than those with ALAD1-1. These data provide further evidence that the ALAD genotype modifies the toxicokinetics of lead-for example, by differential binding of current lead stores or by differences in long-term retention and deposition of lead.

Reiko Akagi - One of the best experts on this subject based on the ideXlab platform.

  • co synthesis of human δ aminolevulinate dehydratase ALAD mutants with the wild type enzyme in cell free system critical importance of conformation on enzyme activity
    Journal of Clinical Biochemistry and Nutrition, 2008
    Co-Authors: Rikako Inoue, Reiko Akagi
    Abstract:

    Properties of mutant δ-aminolevulinate dehydratase (ALAD) found in patients with ALAD porphyria were studied by enzymological and immunological analyses after the synthesis of enzyme complexes using a cell-free system. Enzyme activities of homozygous G133R, K59N/G133R, V153M, and E89K mutants were 11%, 22%, 67%, and 75% of the wild-type ALAD, respectively, whereas that of K59N, a normal variant, was 112%. Enzyme activities of L273R, C132R and F12L were undetectable. Co-synthesis of F12L, L273R, G133R, K59N/G133R, or C132R mutants with the wild-type at various ratios showed that ALAD activity was proportionally decreased in the amount of the wild-type in the complex. In contrast, co-synthesis of V153M, K59N, and E89K with the wild-type did not influence enzyme activity of the wild-type. Surface charge changes in K59N, E89K, C132R and G133R predicted by mutations were also confirmed by native polyacrylamide gel electrophoresis. A compound E89K and C132R complex showed ALAD activity similar to that was found in erythrocytes of the patient. These findings indicate that cell-free synthesis of ALAD proteins reflects enzymatic activities found in patients, and suggest that, in addition to the direct effect of mutations on the catalytic activity, conformational effects play an important role in determining enzyme activity.

  • δ aminolevulinate dehydratase ALAD porphyria the first case in north america with two novel ALAD mutations
    Molecular Genetics and Metabolism, 2006
    Co-Authors: Reiko Akagi, Rikako Inoue, Karl E. Anderson, Noriko Kato, Eileen K. Jaffe, Shigeru Sassa
    Abstract:

    Abstract The molecular basis of the enzymatic defect responsible for δ-aminolevulinate dehydratase (ALAD) porphyria (ADP) was investigated in a 14-year-old male who presented clinical and laboratory findings typical of ADP. Nucleotide sequence analysis of ALAD cDNAs from the proband revealed two novel mutations, a 265G to A base transition (C1) and a 394C to T base transition (C2), resulting in amino acid substitutions, Glu89Lys and Cys132Arg, respectively. Both mutations were present within exon 5 of the ALAD gene, and appeared to influence the binding of zinc to the enzyme which is essential for enzyme activity. It was found that the C1 mutation was inherited from his father, while the C2 mutation was from his mother. Expression of these mutant ALAD cDNAs in Chinese hamster ovary cells produced normal ALAD mRNA levels, but markedly decreased ALAD protein and enzyme activity. These results suggest that the combination of the two aberrant ALADs with little enzyme activity accounts for the markedly decreased ALAD activity observed in the proband. This case represents the molecular analysis of the ALAD gene defects in the first case of ADP identified in North America, who is a compound heterozygote for two novel ALAD gene defects.

  • dual gene defects involving δ aminolaevulinate dehydratase and coproporphyrinogen oxidase in a porphyria patient
    British Journal of Haematology, 2006
    Co-Authors: Rikako Inoue, Shikibu Muranaka, Tsuyoshi Tahara, Shigeru Taketani, John D Phillips, Reiko Akagi, Karl E. Anderson, Shigeru Sassa
    Abstract:

    Summary A Caucasian male had symptoms of acute porphyria, with increases in urinary δ-aminolaevulinic acid (ALA), porphobilinogen (PBG) and coproporphyrin that were consistent with hereditary coproporphyria (HCP). However, a greater than expected increase in ALA, compared with PBG, and a substantial increase in erythrocyte zinc protoporphyrin, suggested additional ALA dehydratase (ALAD) deficiency. Nucleotide sequence analysis of coproporphyrinogen oxidase (CPO) cDNA of the patient, but not of the parents, revealed a novel nucleotide transition G835C, resulting in an amino acid change, G279R. The mutant CPO protein expressed in Escherichia coli was unstable, and produced about 5% of activity compared with the wild-type CPO. Erythrocyte ALAD activity was 32% of normal in the proband. Nucleotide sequence analysis of cloned ALAD cDNAs from the patient revealed a C36G base transition (F12L amino acid change). The F12L ALAD mutation, which was found in the mother and a brother, was previously described, and is known to lack any enzyme activity. This patient thus represents the first case of porphyria where both CPO and ALAD deficiencies were demonstrated at the molecular level.

  • δ-Aminolevulinate dehydratase (ALAD) porphyria : The first case in North America with two novel ALAD mutations
    Molecular genetics and metabolism, 2005
    Co-Authors: Reiko Akagi, Rikako Inoue, Karl E. Anderson, Noriko Kato, Eileen K. Jaffe, Shigeru Sassa
    Abstract:

    The molecular basis of the enzymatic defect responsible for delta-aminolevulinate dehydratase (ALAD) porphyria (ADP) was investigated in a 14-year-old male who presented clinical and laboratory findings typical of ADP. Nucleotide sequence analysis of ALAD cDNAs from the proband revealed two novel mutations, a 265G to A base transition (C1) and a 394C to T base transition (C2), resulting in amino acid substitutions, Glu89Lys and Cys132Arg, respectively. Both mutations were present within exon 5 of the ALAD gene, and appeared to influence the binding of zinc to the enzyme which is essential for enzyme activity. It was found that the C1 mutation was inherited from his father, while the C2 mutation was from his mother. Expression of these mutant ALAD cDNAs in Chinese hamster ovary cells produced normal ALAD mRNA levels, but markedly decreased ALAD protein and enzyme activity. These results suggest that the combination of the two aberrant ALADs with little enzyme activity accounts for the markedly decreased ALAD activity observed in the proband. This case represents the molecular analysis of the ALAD gene defects in the first case of ADP identified in North America, who is a compound heterozygote for two novel ALAD gene defects.

  • highly heterogeneous nature of δ aminolevulinate dehydratase ALAD deficiencies in ALAD porphyria
    Blood, 2001
    Co-Authors: Motoyoshi Maruno, Reiko Akagi, Kazumichi Furuyama, Yutaka Horie, Kuniaki Meguro, Luba Garbaczewski, Nicholas Chiorazzi, Manfred O Doss, A Hassoun, R Mercelis
    Abstract:

    The properties of 9 delta -aminolevulinate dehydratase (ALAD) mutants from patients with ALAD porphyria (ADP) were examined by bacterial expression of their complementary DNAs and by enzymologic and immunologic assays. ALADs were expressed as glutathione-S-transferase (GST) fusion proteins in Escherichia coli and purified by glutathione-affinity column chromatography. The GST-ALAD fusion proteins were recognized by anti-ALAD antibodies and were enzymatically active as ALAD. The enzymatic activities of 3 ALAD mutants, K59N, A274T, and V153M, were 69.9%, 19.3%, and 41.0% of that of the wild-type ALAD, respectively, whereas 6 mutants, G133R, K59N/G133R, F12L, R240W, V275M, and deITC, showed little activity (< 8%). These variations generally reflect the phenotype of ALAD in vivo in patients with ADP and indicate that GST-ALAD fusion protein is indeed useful for predicting of the phenotype of ALAD mutants. The location of F12L mutation in the enzyme's molecular structure indicates that its disturbance of the quaternary contact of the ALAD dimer appears to have a significant influence on the enzymatic activity. Mouse monoclonal antibodies to human ALAD were developed that specifically recognized a carboxy terminal portion of ALAD, or other regions In the enzyme. This study represents the first complete analysis of 9 mutants of ALAD identified In ADP and indicates the highly heterogeneous nature of mutations in this disorder. (Blood. 2001;97:2972-2978) (C) 2001 by The American Society of Hematology.

Shigeru Sassa - One of the best experts on this subject based on the ideXlab platform.

  • δ aminolevulinate dehydratase ALAD porphyria the first case in north america with two novel ALAD mutations
    Molecular Genetics and Metabolism, 2006
    Co-Authors: Reiko Akagi, Rikako Inoue, Karl E. Anderson, Noriko Kato, Eileen K. Jaffe, Shigeru Sassa
    Abstract:

    Abstract The molecular basis of the enzymatic defect responsible for δ-aminolevulinate dehydratase (ALAD) porphyria (ADP) was investigated in a 14-year-old male who presented clinical and laboratory findings typical of ADP. Nucleotide sequence analysis of ALAD cDNAs from the proband revealed two novel mutations, a 265G to A base transition (C1) and a 394C to T base transition (C2), resulting in amino acid substitutions, Glu89Lys and Cys132Arg, respectively. Both mutations were present within exon 5 of the ALAD gene, and appeared to influence the binding of zinc to the enzyme which is essential for enzyme activity. It was found that the C1 mutation was inherited from his father, while the C2 mutation was from his mother. Expression of these mutant ALAD cDNAs in Chinese hamster ovary cells produced normal ALAD mRNA levels, but markedly decreased ALAD protein and enzyme activity. These results suggest that the combination of the two aberrant ALADs with little enzyme activity accounts for the markedly decreased ALAD activity observed in the proband. This case represents the molecular analysis of the ALAD gene defects in the first case of ADP identified in North America, who is a compound heterozygote for two novel ALAD gene defects.

  • dual gene defects involving δ aminolaevulinate dehydratase and coproporphyrinogen oxidase in a porphyria patient
    British Journal of Haematology, 2006
    Co-Authors: Rikako Inoue, Shikibu Muranaka, Tsuyoshi Tahara, Shigeru Taketani, John D Phillips, Reiko Akagi, Karl E. Anderson, Shigeru Sassa
    Abstract:

    Summary A Caucasian male had symptoms of acute porphyria, with increases in urinary δ-aminolaevulinic acid (ALA), porphobilinogen (PBG) and coproporphyrin that were consistent with hereditary coproporphyria (HCP). However, a greater than expected increase in ALA, compared with PBG, and a substantial increase in erythrocyte zinc protoporphyrin, suggested additional ALA dehydratase (ALAD) deficiency. Nucleotide sequence analysis of coproporphyrinogen oxidase (CPO) cDNA of the patient, but not of the parents, revealed a novel nucleotide transition G835C, resulting in an amino acid change, G279R. The mutant CPO protein expressed in Escherichia coli was unstable, and produced about 5% of activity compared with the wild-type CPO. Erythrocyte ALAD activity was 32% of normal in the proband. Nucleotide sequence analysis of cloned ALAD cDNAs from the patient revealed a C36G base transition (F12L amino acid change). The F12L ALAD mutation, which was found in the mother and a brother, was previously described, and is known to lack any enzyme activity. This patient thus represents the first case of porphyria where both CPO and ALAD deficiencies were demonstrated at the molecular level.

  • δ-Aminolevulinate dehydratase (ALAD) porphyria : The first case in North America with two novel ALAD mutations
    Molecular genetics and metabolism, 2005
    Co-Authors: Reiko Akagi, Rikako Inoue, Karl E. Anderson, Noriko Kato, Eileen K. Jaffe, Shigeru Sassa
    Abstract:

    The molecular basis of the enzymatic defect responsible for delta-aminolevulinate dehydratase (ALAD) porphyria (ADP) was investigated in a 14-year-old male who presented clinical and laboratory findings typical of ADP. Nucleotide sequence analysis of ALAD cDNAs from the proband revealed two novel mutations, a 265G to A base transition (C1) and a 394C to T base transition (C2), resulting in amino acid substitutions, Glu89Lys and Cys132Arg, respectively. Both mutations were present within exon 5 of the ALAD gene, and appeared to influence the binding of zinc to the enzyme which is essential for enzyme activity. It was found that the C1 mutation was inherited from his father, while the C2 mutation was from his mother. Expression of these mutant ALAD cDNAs in Chinese hamster ovary cells produced normal ALAD mRNA levels, but markedly decreased ALAD protein and enzyme activity. These results suggest that the combination of the two aberrant ALADs with little enzyme activity accounts for the markedly decreased ALAD activity observed in the proband. This case represents the molecular analysis of the ALAD gene defects in the first case of ADP identified in North America, who is a compound heterozygote for two novel ALAD gene defects.

  • a novel mutation of δ aminolaevulinate dehydratase in a healthy child with 12 erythrocyte enzyme activity
    British Journal of Haematology, 1999
    Co-Authors: Reiko Akagi, Yumiko Yasui, Pauline Harper, Shigeru Sassa
    Abstract:

    Cloning, expression and phenotype studies of the defective gene for δ-aminolaevulinate dehydratase (ALAD) in a family with an asymptomatic girl who had ALAD deficiency were carried out. The proband was identified by neonatal ALAD screening, and had erythrocyte ALAD activity at 12% of the normal control. She was heterozygous for ALAD deficiency, which was inherited from her father. Nucleotide sequence analysis of the cloned ALAD cDNA revealed C36 to G and T168 to C mutations on the same allele. The former mutation resulted in F12L substitution, whereas the latter was a silent mutation. All family members who had decreased ALAD activity had the same mutation. Expression of the mutant ALAD cDNA in Chinese hamster ovary cells produced an ALAD protein without significant enzyme activity. Additionally, the mutant ALAD cDNA which encodes F12L substitution produced an aberrant migration pattern in polyacrylamide gel electrophoresis under denaturing conditions. This finding probably reflects an abnormal folding of the F12L protein, since the mutation occurred in the α1 helix of the N-terminal arm of the enzyme, which is involved in the extensive quaternary interactions among the subunits. This is also the first report of ALAD gene mutation in an asymptomatic subject.

  • Lead Binding to δ-Aminolevulinic Acid Dehydratase (ALAD) in Human Erythrocytes
    Pharmacology & toxicology, 1997
    Co-Authors: Ingvar A. Bergdahl, Shigeru Sassa, Anders Grubb, Andrejs Schütz, Robert J. Desnick, James G. Wetmur, Staffan Skerfving
    Abstract:

    Over 99% of the lead present in blood is usually found in erythrocytes. To investigate the nature of this selective accumulation of lead in erythrocytes, the specific binding of lead to proteins in human erythrocytes was studied using liquid chromatography coupled to inductively coupled plasma mass spectrometry (LC-ICP-MS). The principal lead-binding protein had a mass of approximately 240 kDa, and adsorption to specific antibodies showed that protein was delta-aminolevulinic acid dehydratase (ALAD). Thus, the previous notion that lead in erythrocytes was bound primarily to haemoglobin has to be revised. Furthermore, in lead-exposed workers, the percentage of lead bound to ALAD was influenced by a common polymorphism in the ALAD gene. Specifically, in seven carriers of the ALAD2 allele, 84% of the protein-bound lead recovered was bound to ALAD compared to 81% in seven homozygotes for the ALAD1 allele whose erythrocytes were matched for blood-lead concentration. The small difference was statistically significant in Wilcoxon matched-pairs signed-rank test (P = 0.03). No ALAD allele-specific difference in ALAD-bound lead was found among 20 unexposed controls. Perhaps the difference in ALAD-bound lead can provide an explanation for the previously reported finding of higher blood-lead levels among carriers of the ALAD2 allele than among ALAD1 homozygotes in lead-exposed populations.

Rikako Inoue - One of the best experts on this subject based on the ideXlab platform.

  • co synthesis of human δ aminolevulinate dehydratase ALAD mutants with the wild type enzyme in cell free system critical importance of conformation on enzyme activity
    Journal of Clinical Biochemistry and Nutrition, 2008
    Co-Authors: Rikako Inoue, Reiko Akagi
    Abstract:

    Properties of mutant δ-aminolevulinate dehydratase (ALAD) found in patients with ALAD porphyria were studied by enzymological and immunological analyses after the synthesis of enzyme complexes using a cell-free system. Enzyme activities of homozygous G133R, K59N/G133R, V153M, and E89K mutants were 11%, 22%, 67%, and 75% of the wild-type ALAD, respectively, whereas that of K59N, a normal variant, was 112%. Enzyme activities of L273R, C132R and F12L were undetectable. Co-synthesis of F12L, L273R, G133R, K59N/G133R, or C132R mutants with the wild-type at various ratios showed that ALAD activity was proportionally decreased in the amount of the wild-type in the complex. In contrast, co-synthesis of V153M, K59N, and E89K with the wild-type did not influence enzyme activity of the wild-type. Surface charge changes in K59N, E89K, C132R and G133R predicted by mutations were also confirmed by native polyacrylamide gel electrophoresis. A compound E89K and C132R complex showed ALAD activity similar to that was found in erythrocytes of the patient. These findings indicate that cell-free synthesis of ALAD proteins reflects enzymatic activities found in patients, and suggest that, in addition to the direct effect of mutations on the catalytic activity, conformational effects play an important role in determining enzyme activity.

  • δ aminolevulinate dehydratase ALAD porphyria the first case in north america with two novel ALAD mutations
    Molecular Genetics and Metabolism, 2006
    Co-Authors: Reiko Akagi, Rikako Inoue, Karl E. Anderson, Noriko Kato, Eileen K. Jaffe, Shigeru Sassa
    Abstract:

    Abstract The molecular basis of the enzymatic defect responsible for δ-aminolevulinate dehydratase (ALAD) porphyria (ADP) was investigated in a 14-year-old male who presented clinical and laboratory findings typical of ADP. Nucleotide sequence analysis of ALAD cDNAs from the proband revealed two novel mutations, a 265G to A base transition (C1) and a 394C to T base transition (C2), resulting in amino acid substitutions, Glu89Lys and Cys132Arg, respectively. Both mutations were present within exon 5 of the ALAD gene, and appeared to influence the binding of zinc to the enzyme which is essential for enzyme activity. It was found that the C1 mutation was inherited from his father, while the C2 mutation was from his mother. Expression of these mutant ALAD cDNAs in Chinese hamster ovary cells produced normal ALAD mRNA levels, but markedly decreased ALAD protein and enzyme activity. These results suggest that the combination of the two aberrant ALADs with little enzyme activity accounts for the markedly decreased ALAD activity observed in the proband. This case represents the molecular analysis of the ALAD gene defects in the first case of ADP identified in North America, who is a compound heterozygote for two novel ALAD gene defects.

  • dual gene defects involving δ aminolaevulinate dehydratase and coproporphyrinogen oxidase in a porphyria patient
    British Journal of Haematology, 2006
    Co-Authors: Rikako Inoue, Shikibu Muranaka, Tsuyoshi Tahara, Shigeru Taketani, John D Phillips, Reiko Akagi, Karl E. Anderson, Shigeru Sassa
    Abstract:

    Summary A Caucasian male had symptoms of acute porphyria, with increases in urinary δ-aminolaevulinic acid (ALA), porphobilinogen (PBG) and coproporphyrin that were consistent with hereditary coproporphyria (HCP). However, a greater than expected increase in ALA, compared with PBG, and a substantial increase in erythrocyte zinc protoporphyrin, suggested additional ALA dehydratase (ALAD) deficiency. Nucleotide sequence analysis of coproporphyrinogen oxidase (CPO) cDNA of the patient, but not of the parents, revealed a novel nucleotide transition G835C, resulting in an amino acid change, G279R. The mutant CPO protein expressed in Escherichia coli was unstable, and produced about 5% of activity compared with the wild-type CPO. Erythrocyte ALAD activity was 32% of normal in the proband. Nucleotide sequence analysis of cloned ALAD cDNAs from the patient revealed a C36G base transition (F12L amino acid change). The F12L ALAD mutation, which was found in the mother and a brother, was previously described, and is known to lack any enzyme activity. This patient thus represents the first case of porphyria where both CPO and ALAD deficiencies were demonstrated at the molecular level.

  • δ-Aminolevulinate dehydratase (ALAD) porphyria : The first case in North America with two novel ALAD mutations
    Molecular genetics and metabolism, 2005
    Co-Authors: Reiko Akagi, Rikako Inoue, Karl E. Anderson, Noriko Kato, Eileen K. Jaffe, Shigeru Sassa
    Abstract:

    The molecular basis of the enzymatic defect responsible for delta-aminolevulinate dehydratase (ALAD) porphyria (ADP) was investigated in a 14-year-old male who presented clinical and laboratory findings typical of ADP. Nucleotide sequence analysis of ALAD cDNAs from the proband revealed two novel mutations, a 265G to A base transition (C1) and a 394C to T base transition (C2), resulting in amino acid substitutions, Glu89Lys and Cys132Arg, respectively. Both mutations were present within exon 5 of the ALAD gene, and appeared to influence the binding of zinc to the enzyme which is essential for enzyme activity. It was found that the C1 mutation was inherited from his father, while the C2 mutation was from his mother. Expression of these mutant ALAD cDNAs in Chinese hamster ovary cells produced normal ALAD mRNA levels, but markedly decreased ALAD protein and enzyme activity. These results suggest that the combination of the two aberrant ALADs with little enzyme activity accounts for the markedly decreased ALAD activity observed in the proband. This case represents the molecular analysis of the ALAD gene defects in the first case of ADP identified in North America, who is a compound heterozygote for two novel ALAD gene defects.

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  • delta aminolevulinic acid dehydratase genotype modifies four hour urinary lead excretion after oral administration of dimercaptosuccinic acid
    Occupational and Environmental Medicine, 1997
    Co-Authors: Brian S Schwartz, Walter F Stewart, Pornchai Sithisarankul, Paul Timothy Strickland, Karl T Kelsey
    Abstract:

    OBJECTIVES: Previous research suggests that binding of lead by delta-aminolevulinic acid dehydratase (ALAD) may vary by ALAD genotype. This hypothesis was tested by examining whether ALAD genotype modifies urinary lead excretion (DMSA chelatable lead) after oral administration of dimercaptosuccinic acid (DMSA). METHODS: 57 South Korean lead battery manufacturing workers were given 5 mg/kg oral DMSA and urine was collected for four hours. Male workers were randomly selected from two ALAD genotype strata (ALAD1-1, ALAD1-2) from among all current workers in the two plants (n = 290). Subjects with ALAD1-1 (n = 38) were frequency matched with subjects with ALAD1-2 (n = 19) on duration of employment in the lead industry. Blood lead, zinc protoporphyrin, and plasma aminolevulinic acid concentrations, as well as ALAD genotype, duration of exposure, current tobacco use, and weight were examined as predictors or effect modifiers of levels of DMSA chelatable lead. RESULTS: Blood lead concentrations ranged from 11 to 53 micrograms/dl, with a mean (SD) of 25.4 (10.2) micrograms/dl. After 5 mg/kg DMSA orally, the workers excreted a mean (SD) 85.4 (45.0) micrograms lead during a four hour urine collection (range 16.5-184.1 micrograms). After controlling for blood lead concentrations, duration of exposure, current tobacco use, and body weight, subjects with ALAD1-2 excreted, on average, 24 micrograms less lead during the four hour urine collection than did subjects with ALAD1-1 (P = 0.05). ALAD genotype seemed to modify the relation between plasma delta-aminolevulinic acid (ALA) and DMSA chelatable lead. Workers with ALAD1-2 excreted more lead, after being given DMSA, with increasing plasma ALA than did workers with ALAD1-1 (P value for interaction = 0.01). CONCLUSIONS: DMSA chelatable lead may partly reflect the stores of bioavailable lead, and the current data indicate that subjects with ALAD1-2 have lower stores than those with ALAD1-1. These data provide further evidence that the ALAD genotype modifies the toxicokinetics of lead-for example, by differential binding of current lead stores or by differences in long-term retention and deposition of lead.

  • associations of subtypes of hemoglobin with delta aminolevulinic acid dehydratase genotype and dimercaptosuccinic acid chelatable lead levels
    Archives of Environmental Health, 1997
    Co-Authors: Brian S Schwartz, Walter F Stewart, Karl T Kelsey, Joseph Bressler
    Abstract:

    Abstract Hemoglobin in erythrocytes may be an important intravascular site of lead binding. We examined associations of hemoglobin subtypes A1 and A2 with δ-aminolevulinic acid dehydratase (ALAD) genotype, a protein that is another important site of erythrocyte lead binding. After oral administration of dimercaptosuccinic acid (DMSA-chelatable lead), we also examined 4-h lead excretion, which provides an estimate of bioavailable lead stores. We randomly selected 57 South Korean current lead battery manufacturing workers from two plants (N = 290 employees) and from two ALAD genotype strata (ALAD1–1 and ALAD1–2). These workers voluntarily administered 5 mg/kg oral DMSA. We frequency-matched subjects with ALAD1–1 (n = 38) to subjects with ALAD1–2 (n = 19) with respect to duration of employment in the lead industry. Blood lead levels ranged from 11 to 53 μg/dl (mean ± standard deviation, 25.4 ± 10.2 μg/dl). After administration of oral DMSA, workers excreted a mean lead level of 85.4 (standard deviation, 45.0...

  • associations of δ aminolevulinic acid dehydratase genotype with plant exposure duration and blood lead and zinc protoporphyrin levels in korean lead workers
    American Journal of Epidemiology, 1995
    Co-Authors: Brian S Schwartz, Walter F Stewart, Byung Kook Lee, Kyu Dong Ahn, Kathryn Springer, Karl T Kelsey
    Abstract:

    Previous studies have suggested that polymorphisms in δ-aminolevulinic acid dehydratase (ALAD), a heme synthetic enzyme, may be associated with differences in blood lead levels, perhaps due to differential binding of lead in erythrocytes. The authors examined associations of ALAD genotype with blood lead and zinc protoporphyrin (ZPP) levels, exposure duration, sex, and plant in 308 currently exposed lead workers in three lead storage battery manufacturing plants in the Republic of Korea in 1993. The overall prevalence of the variant allele, ALAD 2 , was 11%, but prevalence varied by plant (p = 0.02 : 8.6% in plant A, 20% in plant B, and 22% in plant C). While ALAD 2 was not associated with mean blood lead levels, the allele was associated with blood lead levels greater than or equal to 40 μg/dl (crude odds ratio (OR) = 2.6, 95% confidence interval (CI) 1.1-6.3 ; adjusted OR = 2.3, 95% CI 0.8-6.2, with adjustment for sex, plant, and exposure duration) and with exposure durations greater than 6 years (adjusted OR = 2.5, 95% CI 1.2-5.4, with adjustment for blood lead, sex, and plant). Among workers in plant C, the highest exposure plant, ALAD 2 was associated with lower ZPP levels when controlling for blood lead levels. These data suggest that lead toxicokinetics may be modified by ALAD genotype and that ALAD 2 may be protective for the health effects of lead. ALAD genotype also appears to have been a selection factor for current lead exposure status in the studied workers. Am J Epidemiol 1995 ; 142 :738-45.