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B R Akerman

Publications and source records attributed to B R Akerman.

18 recordsLinked to original sources
PubMed2001

In vivo variability of TMA oxidation is partially mediated by polymorphisms of the FMO3 gene.

Trimethylaminuria (TMAU) results from an accumulation of an excessive amount of unoxidized trimethylamine that is excreted in urine and body secretions. Mutations of the flavin-containing monooxygenase 3 (FMO3) gene (a hepatic phase I drug-metabolizing enzyme) account for the severe recessively encoded form of this condition. We have previously described a number of FMO3 polymorphisms which in vitro exhibit reduced substrate affinity for several FMO substrates. Here we show that three prevalent polymorphisms (E158K, V257M, and E308G) inherited in particular combinations confer a slight decrease in TMA oxidation under normal physiological conditions, which may be clinically "silent." With the use of substrate loading or with the interaction of other known modulators of FMO3 activity such as hormonal influences, these genotypes may predispose to mild TMAU.

Alleles↗
PubMed2001

A novel deletion in the flavin-containing monooxygenase gene (FMO3) in a Greek patient with trimethylaminuria.

Mutations of the flavin-containing monooxygenase type 3 gene (FMO3) that encode the major functional form present in adult human liver, have been shown to cause trimethylaminuria. We now report a novel homozygous deletion of exons 1 and 2 in an Australian of Greek ancestry with TMAuria, the first report of a deletion causative of trimethylaminuria. The deletion occurs 328 bp upstream from exon 1. The 3'-end of the deletion occurs in intron 2, 10013 base pairs downstream from the end of exon 2. The deletion is 12226 bp long. For the proband homozygous for the human FMO3 gene deletion, it is predicted that in addition to loss of monooxygenase function for human FMO3 substrates, such as TMA and other amines, the proband will exhibit decreased tolerance of biogenic amines, both medicinal and those found in foods.

Adolescent↗
PubMed2000

Characterization of phenylketonuria missense substitutions, distant from the phenylalanine hydroxylase active site, illustrates a paradigm for mechanism and potential modulation of phenotype.

Missense mutations account for 48% of all reported human disease-causing alleles. Since few are predicted to ablate directly an enzyme's catalytic site or other functionally important amino acid residues, how do most missense mutations cause loss of function and lead to disease? The classic monogenic phenotype hyperphenylalaninemia (HPA), manifesting notably as phenylketonuria (PKU), where missense mutations in the PAH gene compose 60% of the alleles impairing phenylalanine hydroxylase (PAH) function, allows us to examine this question. Here we characterize four PKU-associated PAH mutations (F39L, K42I, L48S, I65T), each changing an amino acid distant from the enzyme active site. Using three complementary in vitro protein expression systems, and 3D-structural localization, we demonstrate a common mechanism. PAH protein folding is affected, causing altered oligomerization and accelerated proteolytic degradation, leading to reduced cellular levels of this cytosolic protein. Enzyme specific activity and kinetic properties are not adversely affected, implying that the only way these mutations reduce enzyme activity within cells in vivo is by producing structural changes which provoke the cell to destroy the aberrant protein. The F39L, L48S, and I65T PAH mutations were selected because each is associated with a spectrum of in vivo HPA among patients. Our in vitro data suggest that interindividual differences in cellular handling of the mutant, but active, PAH proteins will contribute to the observed variability of phenotypic severity. PKU thus supports a newly emerging paradigm both for mechanism whereby missense mutations cause genetic disease and for potential modulation of a disease phenotype.

ATP-Binding Cassette Transporters↗
PubMed2000

Population-specific polymorphisms of the human FMO3 gene: significance for detoxication.

Flavin-containing monooxygenase form 3 (FMO3) is one of the major enzyme systems that protect humans from the potentially toxic properties of drugs and chemicals. FMO3 converts nucleophilic heteroatom-containing chemicals and endogenous materials to polar metabolites, which facilitates their elimination. For example, the tertiary amine trimethylamine is N-oxygenated by human FMO3 to trimethylamine N-oxide, and trimethylamine N-oxide is excreted in a detoxication and deoderation process. In normal humans, virtually all trimethylamine is metabolized to trimethylamine N-oxide. In a few humans, trimethylamine is not efficiently metabolized to trimethylamine N-oxide, and those individuals suffer from trimethylaminuria, or fishlike odor syndrome. Previously, we identified mutations of the FMO3 gene that cause trimethylaminuria. We now report two prevalent polymorphisms of this gene (K158E and V257M) that modulate the activity of human FMO3. These polymorphisms are widely distributed in Canadian and Australian white populations. In vitro analysis of wild-type and variant human FMO3 proteins expressed from the cDNA for the two naturally occurring polymorphisms showed differences in substrate affinities for nitrogen-containing substrates. Thus, for polymorphic forms of human FMO3, lower k(cat)/K(m) values for N-oxygenation of 10-(N, N-dimethylaminopentyl)-2-(trifluoromethyl) phenothiazine, trimethylamine, and tyramine were observed. On the basis of in vitro kinetic parameters, human FMO1 does not significantly contribute to human metabolism of trimethylamine or tyramine. The results imply that prevalent polymorphisms of the human FMO3 gene may contribute to low penetrance predispositions to diseases associated with adverse environmental exposures to heteroatom-containing chemicals, drugs, and endogenous amines.

Australia↗
PubMed1999

Two novel mutations of the FMO3 gene in a proband with trimethylaminuria.

The mammalian flavin-containing monooxygenases catalyze the NADPH-dependent N-oxygenation of nucleophilic nitrogen-, sulfur-, and phosphorus-containing chemicals, drugs, and xenobiotics, including trimethylamine. The FMO3 gene encodes the dominant catalytically active isoform present in human liver. We have identified two missense mutations in the coding region of the gene in a proband with trimethylaminuria (TMA): M66I and R492W. Whereas two mutations (P153L, E305X) accounted for TMA in our eight unrelated previously documented Australian families of British origin, the present report is the first evidence of compound heterozygosity for two rare mutations in a proband with this disorder. This suggests that other rarer alleles, also causing TMA, will be found in the same populations.

Adult↗
PubMed1999

Trimethylaminuria is caused by mutations of the FMO3 gene in a North American cohort.

Trimethylaminuria (TMAuria) (McKusick 602079) first described in 1970 is an autosomal recessive condition caused by a partial or total incapacity to catalyze the N-oxygenation of the odorous compound trimethylamine (TMA). The result is a severe body odor and associated psychosocial conditions. This inborn error of metabolism, previously thought to be rare, is now being increasingly detected in severe and milder presentations. Mutations of a phase 1 detoxicating gene, flavin-containing monooxygenase 3 (FMO3), have been shown to cause TMAuria. Herein we describe a cohort of individuals ascertained in North America with severe TMAuria, defined by a reduction of TMA oxidation below 50% of normal with genotype-phenotype correlations. We detected four new FMO3 mutations; two were missense (A52T and R387L), one was nonsense (E314X). The fourth allele is apparently composed of two relatively common polymorphisms (K158-G308) found in the general population. On the basis of this study we conclude that one common mutation and an increasing number of private mutations in individuals of different ethnic origins cause TMAuria in this cohort.

Adult↗
PubMed1998

Mutations of the flavin-containing monooxygenase gene (FMO3) cause trimethylaminuria, a defect in detoxication.

Individuals with the recessive condition trimethylaminuria exhibit variation in metabolic detoxication of xenobiotics by hepatic flavin-containing monooxygenases. We show here that mutations in the human flavin-containing monooxygenase isoform 3 gene ( FMO3 ) impair N -oxygenation of xenobiotics and are responsible for the trimethylaminuria phenotype. Three disease-causing mutations in nine Australian-born probands have been identified which share a particular polymorphic haplotype. Nonsense and missense mutations are associated with a severe phenotype and are also implicated in impaired metabolism of other nitrogen- and sulfur-containing substrates including biogenic amines, both clinically and when mutated proteins expressed from cDNA are studied in vitro . These findings illustrate the critical role played by human FMO3 in the metabolism of xenobiotic substrates and endogenous amines.

Adolescent↗
PubMed1997

Novel mutations and DNA-based screening in non-Jewish carriers of Tay-Sachs disease.

We have evaluated the feasibility of using PCR-based mutation screening for non-Jewish enzyme-defined carriers identified through Tay-Sachs disease-prevention programs. Although Tay-Sachs mutations are rare in the general population, non-Jewish individuals may be screened as spouses of Jewish carriers or as relatives of probands. In order to define a panel of alleles that might account for the majority of mutations in non-Jewish carriers, we investigated 26 independent alleles from 20 obligate carriers and 3 affected individuals. Eighteen alleles were represented by 12 previously identified mutations, 7 that were newly identified, and 1 that remains unidentified. We then investigated 46 enzyme-defined carrier alleles: 19 were pseudodeficiency alleles, and five mutations accounted for 15 other alleles. An eighth new mutation was detected among enzyme-defined carriers. Eleven alleles remain unidentified, despite the testing for 23 alleles. Some may represent false positives for the enzyme test. Our results indicate that predominant mutations, other than the two pseudodeficiency alleles (739C-->T and 745C-->T) and one disease allele (IVS9+1G-->A), do not occur in the general population. This suggests that it is not possible to define a collection of mutations that could identify an overwhelming majority of the alleles in non-Jews who may require Tay-Sachs carrier screening. We conclude that determination of carrier status by DNA analysis alone is inefficient because of the large proportion of rare alleles. Notwithstanding the possibility of false positives inherent to enzyme screening, this method remains an essential component of carrier screening in non-Jews. DNA screening can be best used as an adjunct to enzyme testing to exclude known HEXA pseudodeficiency alleles, the IVS9+1G-->A disease allele, and other mutations relevant to the subject's genetic heritage.

California↗
PubMed1996

Dramatically different phenotypes in mouse models of human Tay-Sachs and Sandhoff diseases.

We have generated mouse models of human Tay-Sachs and Sandhoff diseases by targeted disruption of the Hexa (alpha subunit) or Hexb (beta subunit) genes, respectively, encoding lysosomal beta-hexosaminidase A (structure, alpha) and B (structure, beta beta). Both mutant mice accumulate GM2 ganglioside in brain, much more so in Hexb -/- mice, and the latter also accumulate glycolipid GA2. Hexa -/- mice suffer no obvious behavioral or neurological deficit, while Hexb -/- mice develop a fatal neurodegenerative disease, with spasticity, muscle weakness, rigidity, tremor and ataxia. The Hexb -/- but not the Hexa -/- mice have massive depletion of spinal cord axons as an apparent consequence of neuronal storage of GM2. We propose that Hexa -/- mice escape disease through partial catabolism of accumulated GM2 via GA2 (asialo-GM2) through the combined action of sialidase and beta-hexosaminidase B.

Animals↗
PubMed1994

Mutations participating in interallelic complementation in propionic acidemia.

Deficiency of propionyl-CoA carboxylase (PCC; alpha 4 beta 4) results in the rare, autosomal recessive disease propionic acidemia. Cell fusion experiments have revealed two complementation groups, pccA and pccB, corresponding to defects of the PCCA (alpha-subunit) and PCCB (beta-subunit) genes, respectively. The pccBCC group includes subgroups, pccB and pccC, which are thought to reflect interallelic complementation between certain mutations of the PCCB gene. In this study, we have identified the mutations in two pccB, one pccC, and two pccBC cell lines and have deduced those alleles participating in interallelic complementation. One pccB line was a compound heterozygote of Pro228Leu and Asn536Asp. The latter mutation was also detected in a noncomplementing pccBC line. This leaves Pro228Leu responsible for complementation in the pccB cells. The second pccB line contained an insertional duplication, dupKICK140-143, and a splice mutation IVS + 1 G-->T, located after Lys466. We suggest that the dupKICK mutation is the complementing allele, since the second allele is incompatible with normal splicing. The pccC line studied was homozygous for Arg410Trp, which is necessarily the complementing allele in that line. For a second pccC line, we previously had proposed that delta Ile408 was the complementing allele. We now show that its second allele, "Ins.Del," a 14-bp deletion replaced by a 12-bp insertion beginning at codon 407, fails to complement in homozygous form. We conclude that the interallelic complementation results from mutations in domains that can interact between beta-subunits in the PCC heteromer to restore enzymatic function. On the basis of sequence homology with the Propionibacterium shermanii transcarboxylase 12S subunit, we suggest that the pccC domain, defined by Ile408 and Arg410, may involve the propionyl-CoA binding site.

Alleles↗
PubMed1992

Molecular basis of hexosaminidase A deficiency and pseudodeficiency in the Berks County Pennsylvania Dutch.

Following the birth of two infants with Tay-Sachs disease (TSD), a non-Jewish, Pennsylvania Dutch kindred was screened for TSD carriers using the biochemical assay. A high frequency of individuals who appeared to be TSD heterozygotes was detected (Kelly et al., 1975). Clinical and biochemical evidence suggested that the increased carrier frequency was due to at least two altered alleles for the hexosaminidase A alpha-subunit. We now report two mutant alleles in this Pennsylvania Dutch kindred, and one polymorphism. One allele, reported originally in a French TSD patient (Akli et al., 1991), is a GT-->AT transition at the donor splice-site of intron 9. The second, a C-->T transition at nucleotide 739 (Arg247Trp), has been shown by Triggs-Raine et al. (1992) to be a clinically benign "pseudodeficient" allele associated with reduced enzyme activity against artificial substrate. Finally, a polymorphism [G-->A (759)], which leaves valine at codon 253 unchanged, is described.

Alleles↗
PubMed1992

A mutation common in non-Jewish Tay-Sachs disease: frequency and RNA studies.

Tay-Sachs disease (TSD) is an autosomal recessive genetic disorder resulting from mutation of the HEXA gene encoding the alpha-subunit of the lysosomal enzyme, beta-N-acetylhexosaminidase A (Hex A). We have discovered that a Tay-Sachs mutation, IVS-9 + 1 G-->A, first detected by Akli et al. (Genomics 11:124-134, 1991), is a common disease allele in non-Jewish Caucasians (10/58 alleles examined). A PCR-based diagnostic test, which detects an NlaIII site generated by the mutation, revealed a frequency among enzyme-defined carriers of 9/64 (14%). Most of those carrying the allele trace their origins to the United Kingdom, Ireland, or Western Europe. It was not identified among 12 Black American TSD alleles or in any of 18 Ashkenazi Jewish, enzyme-defined carriers who did not carry any of the mutations common to this population. No normally spliced RNA was detected in PCR products generated from reverse transcription of RNA carrying the IVS-9 mutation. Instead, the low levels of mRNA from this allele were comprised of aberrant species resulting from the use of either of two cryptic donor sites, one truncating exon 9 and the other within IVS-9, spliced to exon 10. Numerous additional splice products were detected, most involving skipping of one or more surrounding exons. Together with a recently identified allele responsible for Hex A pseudodeficiency (Triggs-Raine et al. Am J Hum Genet, 1992), these two alleles accounted for almost 50% (29/64) of TSD or carrier alleles ascertained by enzyme screening tests in non-Jewish Caucasians.

Alleles↗
PubMed1992

A pseudodeficiency allele common in non-Jewish Tay-Sachs carriers: implications for carrier screening.

Deficiency of beta-hexosaminidase A (Hex A) activity typically results in Tay-Sachs disease. However, healthy subjects found to be deficient in Hex A activity (i.e., pseudodeficient) by means of in vitro biochemical tests have been described. We analyzed the HEXA gene of one pseudodeficient subject and identified both a C739-to-T substitution that changes Arg247----Trp on one allele and a previously identified Tay-Sachs disease mutation on the second allele. Six additional pseudodeficient subjects were found to have the C739-to-T mutation. This allele accounted for 32% (20/62) of non-Jewish enzyme-defined Tay-Sachs disease carriers but for none of 36 Jewish enzyme-defined carriers who did not have one of three known mutations common to this group. The C739-to-T allele, together with a "true" Tay-Sachs disease allele, causes Hex A pseudodeficiency. Given both the large proportion of non-Jewish carriers with this allele and that standard biochemical screening cannot differentiate between heterozygotes for the C739-to-T mutations and Tay-Sachs disease carriers, DNA testing for this mutation in at-risk couples is essential. This could prevent unnecessary or incorrect prenatal diagnoses.

Amino Acid Sequence↗
PubMed1991

Sequence of DNA flanking the exons of the HEXA gene, and identification of mutations in Tay-Sachs disease.

The rapid identification of mutations causing Tay-Sachs disease requires the capacity to readily screen the regions of the HEXA gene most likely to be affected by mutation. We have sequenced the portions of the introns flanking each of the 14 HEXA exons in order to specify oligonucleotide primers for the PCR-dependent amplification of each exon and splice-junction sequence. The amplified products were analyzed, by electrophoresis in nondenaturing polyacrylamide gels, for the presence of either heteroduplexes, derived from the annealing of normal and mutant DNA strands, or single-strand conformational polymorphisms (SSCP), derived from the renaturation of single-stranded DNA. Five novel mutations from Tay-Sachs disease patients were detected: a 5-bp deletion of TCTCC in IVS-9; a 2-bp deletion of TG in exon 5; G78 to A, giving a stop codon in exon 1; G533 to T in exon 5, producing the third amino acid substitution detected at this site; and G to C at position 1 of IVS-2, expected to produce abnormal splicing. In addition, two mutations, (G1496 to A in exon 13 and a 4-bp insertion in exon 11) that have previously been reported were identified.

Alleles↗
PubMed1990

Identification of deletion and triple alpha-globin gene haplotypes in the Montreal beta-thalassemia screening program: implications for genetic medicine.

We obtained blood samples in a screening program designed to detect beta-thalassemia heterozygotes in Montreal; additional samples were obtained from referred persons. We analyzed DNA for variant numbers of alpha-globin genes, notably the alpha-thalassemia2 (-alpha/), alpha-thalassemia1, (- -/), and triplicated alpha-globin gene (alpha alpha alpha/) haplotypes using restriction enzymes and probes for alpha-globin and zeta-globin gene sequences. We estimated the numbers of Montreal residents of Italian and Greek ethnic origin with -alpha/alpha alpha genotype. Thus, 4.3% of Italians and 1.5% of Greeks, or about 7,500 persons, are estimated to be alpha-thalassemia2, trait (silent carriers), largely (80%) in the -alpha 3.7/type I form. The triplicated alpha-globin gene haplotype was also found. The risk of a severe (alpha-thalassemia1) phenotype associated with inheritance of - -/alpha alpha or -alpha/-alpha genotypes was low and was found predominantly in this study, in persons of Asian ethnic origin. The sample of Asians was too small to estimate carrier frequencies; however, based on results from the beta-thalassemia screening program, we estimated that about 4% of Asians (about 1,300 persons) in Montreal are alpha-thalassemia carriers. We identified persons heterozygous for both beta-thalassemia and alpha-thalassemia mutations. In these double heterozygotes, the effect of the triplicated alpha-globin gene was to make the erythrocyte parameters used for screening (MCV and %HbA2) more deviant from normal whereas deletion of 2 alpha-globin genes tended to normalize the erythrocyte values. These findings have implications for the screening program and reproductive counseling.

Adolescent↗