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Biomedical subjects

H G Koch

Publications and source records attributed to H G Koch.

At least 37 records · Page 2Linked to original sources
PubMed2001

Individual blood-brain barrier phenylalanine transport determines clinical outcome in phenylketonuria.

Different clinical outcomes in spite of comparable dietary controls are well known in patients with phenylketonuria. Currently, reasons for this phenomenon are unknown. Kinetic investigations in 15 patients with classic phenylketonuria were performed using in vivo nuclear magnetic resonance spectroscopy before and after an oral phenylalanine load (100 mg/kg body weight). Patients' brain phenylalanine concentrations were quite different in spite of similar blood phenylalanine levels. Interindividual variations of the apparent transport Michaelis constant, K(t,app), covered a range from 0.10 to 1.03 mmol/L. The ratio of the maximal transport velocity, Tmax, over the intracerebral consumption rate, Vmet, varied between 2.61 and 14.0. Both parameters as well as the preload brain phenylalanine levels correlated significantly with the degree of cerebral white matter abnormalities on magnetic resonance images. Correlations of K(t,app), Tmax/Vmet, and the preload brain phenylalanine levels with patients' intelligence scores approached significance. In conclusion, blood-brain barrier phenylalanine transport characteristics and the resultant brain phenylalanine levels seem to be causative factors for the individual clinical outcome in phenylketonuria. This observation may lead to individual dietary recommendations in the future.

Adolescent↗
PubMed2001

Haplotyping of wild type and I278T alleles of the human cystathionine beta-synthase gene based on a cluster of novel SNPs in IVS12.

Homocystinuria is most frequently due to deficiency of cystathionine beta-synthase (CBS). We identified IVS12 as a polymorphism hot spot of the human CBS gene and report five novel single nucleotide polymorphisms (SNPs): g.13514G>A, g.13617A>G, g.13715C>T, g.13800G>A, and g.13904C>T. Analyzing 50 control DNA samples of unaffected and unrelated subjects of German origin the observed frequencies of heterozygosity were 0.02, 0.36, 0.18, 0.36, and 0.36, respectively. These polymorphic markers were combined into four distinct IVS12-haplotypes A1, A2, B1, and B2, revealing frequencies of 0.75, 0.01, 0.15, and 0.09, respectively, with an observed overall frequency of heterozygosity at 0.38. This haplotype system and the SNP c.699 were employed in the analysis of ten alleles affected by the most prevalent CBS mutation, c.833T>C (exon 8; I278T). We found that the I278T alleles segregate with at least two distinct haplotypes characterized by upstream and downstream polymorphic sites instead of sharing a common ancestral haplotype. This was a remarkable finding even in patients with very similar ethnic background. The novel haplotype system may facilitate future studies on the evolution of the CBS gene and might be suited for genotyping of families affected by homocystinuria.

Alleles↗
PubMed2001

Genetic analysis of carbamoylphosphate synthetase I and ornithine transcarbamylase deficiency using fibroblasts.

UNLABELLED: Deficiencies of carbamoylphosphate synthetase or of ornithine transcarbamylase, two urea cycle enzymes located within mitochondria, often present as severe neonatal hyperammonaemic crises and have a poor prognosis. While genetic analysis of the X-chromosomal transmitted ornithine transcarbamylase deficiency (OTC) is performed by exon-wise mutation screening of genomic DNA in most cases, identification of mutations in the autosomal inherited carbamoylphosphate synthetase (CPS 1) deficiency requires analysis of transcripts due to the unknown genomic structure. We tested the hypothesis that CPS 1 and OTC are expressed at low levels in fibroblasts and indeed were able to amplify full-length cDNA from that source. Using a reverse transcriptase polymerase chain reaction based procedure we completely characterised the genetic background in five patients and identified three novel mutations and a novel polymorphism of the CPS 1 gene (deletion/insertion 2170delGCTCinsCCA, nonsense mutation 2359C > T, missense mutation 3161T > G and Thr1406Asn, respectively), as well as the missense mutations 482A > G and 994T > A of the OTC gene. CONCLUSION: Cultured fibroblasts are an easily accessible source for genetic analysis of inborn errors of urea cycle enzymes which are functionally expressed only in liver and gut.

Carbamoyl-Phosphate Synthase (Ammonia)↗
PubMed2001

Protein traffic in bacteria: multiple routes from the ribosome to and across the membrane.

Bacteria use several routes to target their exported proteins to the plasma membrane. The majority are exported through pores formed by SecY and SecE. Two different molecular machineries are used to target proteins to the SecYE translocon. Translocated proteins, synthesized as precursors with cleavable signal sequences, require cytoplasmic chaperones, such as SecB, to remain competent for posttranslational transport. In concert with SecB, SecA targets the precursors to SecY and energizes their translocation by its ATPase activity. The latter function involves a partial insertion of SecA itself into the SecYE translocon, a process that is strongly assisted by a couple of membrane proteins, SecG, SecD, SecF, YajC, and the proton gradient across the membrane. Integral membrane proteins, however, are specifically recognized by a direct interaction between their noncleaved signal anchor sequences and the bacterial signal recognition particle (SRP) consisting of Ffh and 4.5S RNA. Recognition occurs during synthesis at the ribosome and leads to a cotranslational targeting to SecYE that is mediated by FtsY and the hydrolysis of GTP. No other Sec protein is required for integration unless the membrane protein also contains long translocated domains that engage the SecA machinery. Discrimination between SecA/SecB- and SRP-dependent targeting involves the specificity of SRP for hydrophobic signal anchor sequences and the exclusion of SRP from nascent chains of translocated proteins by trigger factor, a ribosome-associated chaperone. The SecYE pore accepts only unfolded proteins. In contrast, a class of redox factor-containing proteins leaves the cell only as completely folded proteins. They are distinguished by a twin arginine motif of their signal sequences that by an unknown mechanism targets them to specific pores. A few membrane proteins insert spontaneously into the bacterial plasma membrane without the need for targeting factors and SecYE. Insertion depends only on hydrophobic interactions between their transmembrane segments and the lipid bilayer and on the transmembrane potential. Finally, outer membrane proteins of Gram-negative bacteria after having crossed the plasma membrane are released into the periplasm, where they undergo distinct folding events until they insert as trimers into the outer membrane. These folding processes require distinct molecular chaperones of the periplasm, such as Skp, SurA, and PpiD.

Adenosine Triphosphatases↗
PubMed2001

Acute and chronic pancreatitis in patients with inborn errors of metabolism.

Acute and chronic recurrent pancreatitis have been reported in patients with a variety of inborn errors of metabolism. Among these are hyperlipidaemias, various disorders of branched-chain amino acid degradation, homocystinuria, haemolytic disorders, acute intermittent porphyria and several amino acid transporter defects. Some of these disease entities are exceedingly rare. In most of these disorders, pancreatitis is not very common and, with the exception of lipoprotein lipase and apolipoprotein C-II deficiency, is neither the leading nor the clinically most distressing manifestation of the underlying metabolic defect. The majority of these syndromes are, however, inherited, and often entire kindreds are carriers of well-defined germline mutations that can, to varying degrees, be associated with pancreatitis. We have reviewed the clinical, biochemical and genetic characteristics of those inborn errors of metabolism because interesting information can be gained from the in regard to the pathophysiology of pancreatitis and because they need to be distinguished from other hereditary causes of the disease.

Acute Disease↗
PubMed2001

Normal clinical outcome in untreated subjects with mild hyperphenylalaninemia.

ABSTRACT There is international consensus that patients with phenylalanine (Phe) levels <360 microM on a free diet do not need Phe-lowering dietary treatment whereas patients with levels >600 microM do. Clinical outcome of patients showing Phe levels between 360 and 600 microM in serum on a free nutrition has so far only been assessed in a small number of cases. Therefore, different recommendations exist for patients with mild hyperphenylalaninemia. We investigated in a nationwide study 31 adolescent and adult patients who persistently displayed serum Phe levels between 360 and 600 microM on a normal nutrition with a corresponding genotype. Because of limited accuracy of measurements, Phe levels should be looked on as an approximation, but not as an absolute limit in every instance. In addition to serum Phe levels, the assessment program consisted of comprehensive psychological testing, magnetic resonance imaging of the head, (1)H magnetic resonance spectroscopy, and genotyping. We found a normal intellectual (intelligence quotient, 103 +/- 15; range, 79-138) and educational (school performance and job career) outcome in these subjects as compared with healthy control subjects (intelligence quotient, 104 +/- 11; range, 80-135). Magnetic resonance imaging revealed no changes of cerebral white matter in any patient, and (1)H magnetic resonance spectroscopy revealed brain Phe levels below the limit of detection (<200 microM). In the absence of any demonstrable effect, dietary treatment is unlikely to be of value in patients with mild hyperphenylalaninemia and serum Phe levels <600 microM on a free nutrition, and should no longer be recommended. Because of a possible late-onset phenylketonuria, Phe levels of untreated patients should be monitored carefully at least during the first year of life. Nevertheless, problems of maternal phenylketonuria should still be taken into account.

Adolescent↗
PubMed2001

High prevalence of the I278T mutation of the human cystathionine beta-synthase detected by a novel screening application.

Classical homocystinuria due to cystathionine beta-synthase deficiency is one of the disorders revealing a high risk of thromboembolic events and vascular disease. This autosomal-recessively inherited metabolic disorder is considered to be rare with an estimated prevalence of 1:130,000 in the German population. In this study, we developed a novel multiplex PCR generating allele specific fragment lengths to analyse individual genotypes of the two most frequent cystathionine beta-synthase alterations, the I278T mutation, which is worldwide found on up to the half of homocystinuric alleles, and the adjacent polymorphism 844ins68. Screening of 200 unrelated German control subjects revealed a frequency of heterozygosity of 1.5% for 1278T corresponding to a calculated frequency of homozygosity of 1:17.800. Our data indicate that homocystinuria due to cystathionine beta-synthase deficiency is a frequently unrecognized disorder resulting in a high risk of thromboembolic events.

Cystathionine beta-Synthase↗
PubMed2000

Allele association studies with SSR and SNP markers at known physical distances within a 1 Mb region embracing the ALDH2 locus in the Japanese, demonstrates linkage disequilibrium extending up to 400 kb.

There has been considerable recent debate concerning the distances over which levels of allelic association useful for genomic quantitative trait locus (QTL) scans can be detected. We have examined simple sequence repeat (SSR) polymorphisms and two single nucleotide polymorphisms (SNPs) in the region flanking the aldehyde dehydrogenase 2 locus, ALDH2, in populations of Japanese alcoholics and controls. These groups differ significantly in the allele frequencies for the functional SNP in exon XII of this gene located on chromosome 12. The results obtained with SSR markers complement recent investigations with SNPs over similar distances at the TCR alpha/delta locus. Significant allelic association with this marker could be detected for SSRs over distances up to 400 kb and over 37 kb for the SNP thereby extending the distance over which LD at this locus could be detected by an order of magnitude. Furthermore, as a proof of principle, we show that comparisons of allele frequency differences for the SSR markers in the case (alcoholics) and control populations would have detected the ALDH2 marker as a putative QTL. Extending the tests to include alleles at two or three flanking loci suggests that the power to detect QTLs through association can be enhanced significantly.

Alcohol Dehydrogenase↗
PubMed2000

SRP-dependent co-translational targeting and SecA-dependent translocation analyzed as individual steps in the export of a bacterial protein.

Recently it has been recognized that the signal recognition particle (SRP) of Escherichia coli represents a specific targeting device for hydrophobic inner membrane proteins. It has remained unclear, however, whether the bacterial SRP functions in concert with SecA, which is required for the translocation of secretory proteins across the inner membrane. Here, we have analyzed a hybrid protein constructed by fusing the signal anchor sequence of an SRP-dependent inner membrane protein (MtlA) to the mature part of an exclusively SecA-requiring secretory protein (OmpA). We show that the signal anchor sequence of MtlA confers the novel properties onto nascent chains of OmpA of being co-translationally recognized and targeted to SecY by SRP. Once targeted to SecY, ribosome-associated nascent chains of the hybrid protein, however, remain untranslocated unless SecA is present. These results indicate that SRP and SecA cooperate in a sequential, non-overlapping manner in the topogenesis of those membrane proteins which, in addition to a signal anchor sequence, harbor a substantial hydrophilic domain to be translocated into the periplasm.

Adenosine Triphosphatases↗
PubMed2000

Dissecting the translocase and integrase functions of the Escherichia coli SecYEG translocon.

Recent evidence suggests that in Escherichia coli, SecA/SecB and signal recognition particle (SRP) are constituents of two different pathways targeting secretory and inner membrane proteins to the SecYEG translocon of the plasma membrane. We now show that a secY mutation, which compromises a functional SecY-SecA interaction, does not impair the SRP-mediated integration of polytopic inner membrane proteins. Furthermore, under conditions in which the translocation of secretory proteins is strictly dependent on SecG for assisting SecA, the absence of SecG still allows polytopic membrane proteins to integrate at the wild-type level. These results indicate that SRP-dependent integration and SecA/SecB-mediated translocation do not only represent two independent protein delivery systems, but also remain mechanistically distinct processes even at the level of the membrane where they engage different domains of SecY and different components of the translocon. In addition, the experimental setup used here enabled us to demonstrate that SRP-dependent integration of a multispanning protein into membrane vesicles leads to a biologically active enzyme.

Adenosine Triphosphatases↗
PubMed2000

Roles of the ccoGHIS gene products in the biogenesis of the cbb(3)-type cytochrome c oxidase.

In many bacteria the ccoGHIS cluster, located immediately downstream of the structural genes (ccoNOQP) of cytochrome cbb(3) oxidase, is required for the biogenesis of this enzyme. Genetic analysis of ccoGHIS in Rhodobacter capsulatus demonstrated that ccoG, ccoH, ccoI and ccoS are expressed independently of each other, and do not form a simple operon. Absence of CcoG, which has putative (4Fe-4S) cluster binding motifs, does not significantly affect cytochrome cbb(3) oxidase activity. However, CcoH and CcoI are required for normal steady-state amounts of the enzyme. CcoI is highly homologous to ATP-dependent metal ion transporters, and appears to be involved in the acquisition of copper for cytochrome cbb(3) oxidase, since a CcoI-minus phenotype could be mimicked by copper ion starvation of a wild-type strain. Remarkably, the small protein CcoS, with a putative single transmembrane span, is essential for the incorporation of the redox-active prosthetic groups (heme b, heme b(3 )and Cu) into the cytochrome cbb(3) oxidase. Thus, the ccoGHIS products are involved in several steps during the maturation of the cytochrome cbb(3) oxidase.

Bacterial Chromatophores↗
PubMed2000

Renal Fanconi syndrome: first sign of partial respiratory chain complex IV deficiency.

A 2-year-old boy who developed hypophosphatemic rickets without signs of muscular weakness or neurological disturbances is presented. Biochemical findings included hypophosphatemia, metabolic acidosis, hypouricemia, hyperphosphaturia, severe glucosuria, generalized hyperaminoaciduria, hypercalciuria, proteinuria with elevated excretion of IgG, transferrin, albumin and high levels of alpha-1-microglobulin. Urine concentration capacity and creatinine clearance were normal. Lactaturia without elevated levels of plasma lactate and a high urinary excretion of beta-hydroxybutyrate were suggestive for mitochondriopathy. Partial deficiency of cytochrome c oxidase (complex IV of the respiratory chain) was found in skeletal muscle. A renal biopsy specimen demonstrated enlarged mitochondria with abnormal arborization and disorientation of the cristae in the proximal tubular cells. Reduced activity of mitochondrial cytochrome c oxidase in tubular cells could be demonstrated by ultracytochemistry. In conclusion, rickets due to the renal Fanconi syndrome can be the first clinical sign of mitochondrial cytopathies without extra-renal symptoms. Elevated excretion of lactate and ketone bodies in urine may serve as a diagnostic marker.

Child, Preschool↗
PubMed2000

Behavioural and emotional problems in early-treated adolescents with phenylketonuria in comparison with diabetic patients and healthy controls.

Even early-treated patients with phenylketonuria (PKU) have a higher risk of psychosocial maladjustment. This study was performed to determine whether emotional and behavioural problems are specific in phenylketonurics and whether they depend on the quality of biochemical control. This comparative study covered 42 PKU patients aged 10-18 years (mean 14.7 years) and 42 diabetic patients matched for sex, age and socioeconomic status. Patients' groups were compared with a control sample of healthy controls (n = 2900) from an epidemiological study. We used the Child Behavior Check List (CBCL) according to Achenbach, intelligence quotient (IQ) test according to Weiss, and monitoring of blood phenylalanine concentrations and HBA1 concentrations. Internalizing problems such as depressive mood, anxiety, physical complaints or social isolation were significantly elevated in both phenylketonuric and diabetic patients, whereas externalizing problems were not. The two patient groups did not differ significantly either in the degree or in the pattern of their psychological profile. In both groups no significant correlations could be computed between the psychological characteristics and the biochemical control, the IQ, and the socioeconomic status. No patient was undergoing psychiatric treatment or psychotherapy. Our results strongly support a psychological perspective for the development of behavioural and emotional problems in both phenylketonuric and diabetic patients. Thus, medical treatment should be accompanied by psychological support for the families.

Adolescent↗
PubMed2000

Genomic structure and transcript variants of the human methylenetetrahydrofolate reductase gene.

The human 5,10-methylenetetrahydrofolate reductase (MTHFR) represents a major enzyme in the folate-dependent regulation of methionine and homocysteine concentrations. Different MTHFR mutations lead either to severe homocystinuria as a multisystem disorder or to moderate hyperhomocysteinaemia, which is a common risk factor for disorders ranging from cardiovasculopathy to spina bifida. The N-terminal part of the human MTHFR gene is incompletely characterised. We report the completed genomic structure of this gene including three novel exonic sequences on the basis of a 5'-RACE and a 4.2 kb cloned fragment of human genomic DNA. We demonstrate the existence of four MTHFR transcripts differing in their first exons. The diversity of transcripts is due to alternative transcription initiation and alternative splicing. Three putative polypeptides of 657, 698, and 680 amino acids are encoded. The novel genomic sequence described here includes putative promoter regions as suggested by the presence of regions homologue to binding sites for SP1, AP1, AP2, CAAT or GC boxes. Furthermore, we provide evidence that there are no TATA-box elements to regulate the human MTHFR gene. The results of our study render the full-length characterisation of affected alleles in severe homocystinuria and moderate hyperhomocysteinaemia due to MTHFR deficiency and provide a basis for investigating the regulation of the human MTHFR gene.

Base Sequence↗
PubMed1999

Lipoprotein (a) and genetic polymorphisms of clotting factor V, prothrombin, and methylenetetrahydrofolate reductase are risk factors of spontaneous ischemic stroke in childhood.

Ischemic stroke is a rare event in childhood. In approximately one third of cases no obvious underlying cause or disorder can be detected. We investigated the importance of genetic risk factors of venous thromboembolism in childhood or stroke in adulthood as risk factors for spontaneous ischemic stroke in children. One hundred forty-eight Caucasian infants and children (aged 0.5 to 16 years) with stroke and 296 age-matched controls from the same geographic areas as the patients were analyzed for increased lipoprotein (a) [Lp(a)] levels >30 mg/dL; for the presence of the factor V (FV) G1691A mutation, the prothrombin (PT) G20210A variant, and the TT677 genotype of methylenetetrahydrofolate reductase (MTHFR); and deficiencies of protein C, protein S, and antithrombin. The following frequencies (patients v controls), odds ratios (ORs), and confidence intervals (CIs) of single risk factors were found: Lp(a) >30 mg/dL (26.4% v 4.7%; OR/CI, 7.2/3.8 to 13.8; P <.0001), FV G1691A (20.2% v 4%; OR/CI, 6/2.97 to 12.1; P <.0001), protein C deficiency (6% v 0.67%; OR/CI, 9.5/2 to 44.6; P =.001), PT G20210A (6% v 1.3%; OR/CI, 4.7/1.4 to 15.6; P =.01), and the MTHFR TT677 genotype (23.6% v 10.4%; OR/CI, 2.4/1.53 to 4.5; P <.0001). A combination of the heterozygous FV G1691A mutation with increased Lp(a) (n = 11) or the MTHFR TT677 genotype (n = 5) was found in 10. 8% of cases, but only 0.3% of controls (OR/CI, 35.75/4.7 to 272; P <. 0001). Increased Lp (a) levels, the FV G1691A mutation, protein C deficiency, the prothrombin G20210A variant, and the MTHFR TT677 are important risk factors for spontaneous ischemic stroke in childhood.

Adolescent↗