Glucose-6-phosphate dehydrogenase (G6PD) iserlohn and G6PD regensburg: Two new severe enzyme defects in german families

Summary Two new inheritable variants of glucose-6-phosphate dehydrogenase have been found in two unrelated German families. Patients with one variant (G6PD Iserlohn, also referred to as G6PD I) suffered from intermittent hemolytic crises caused by fava beans; patients with the other variant (G6PD Regensburg, G6PD II) disclosed chronic nonspherocytic hemolytic anemia aggravated by drug treatment. Due to their unusual biochemical characteristics, the new variants were designated G6PD Iserlohn and G6PD Regensburg. Both variants showed a reduction of enzyme activity to about 6% of the normal in erythrocytes, normal electrophoretic mobility, increased affinity for glucose-6-phosphate, a reduced affinity for NADP and a pH optimum in the neutral region (7.0 and 7.5). G6PD Iserlohn had a decreased affinity for the inhibitor NADPH; G6PD Regensburg had a normal inhibitor constant. Deamino NADP was utilized at an increased rate by G6PD Regensburg. G6PD Iserlohn was thermostable, G6PD Regensburg mildly instable. G6PD activity in leukocytes was normal in G6PD Iserlohn and reduced to the same degree as in erythrocytets in G6PD Regensburg. The cause of the decreased activity of G6PD Iserlohn appears to be in vivo instability; in G6PD Regensburg further mechanisms might include reduced specific activity or reduced synthesis of the variant enzyme.     

Glucose-6-phosphate dehydrogenase (G6PD) mutations and UDP-glucuronosyltransferase promoter polymorphism among G6PD deficient Kuwaitis

Screening of 1,080 Kuwaiti male blood donors for glucose-6-phosphate dehydrogenase (G6PD) deficiency revealed this condition in 70 (6.5%) individuals. Mutation analysis of all 70 G6PD deficient samples performed by PCR/RFLP and direct sequencing identified the 563C-->T (Mediterranean) in 72.9%, 202G-->A (A(-)) in 14.3%, 1003G-->A (Chatham) in 7.1%, and 143T-->C (Aures) in 1.4%. In 3 cases (4.3%) mutations remain unknown. Genotyping of all G6PD deficient samples for UDP-glucuronosyltransferase 1 (UDPGT1) gene promoter polymorphism revealed (ta)6/(ta)6 in 38.6%, (ta)7/(ta)7 in 15.7%, (ta)6/(ta)7 in 44.3%, and (ta)7/(ta)8 allele in 1.4% of cases. Thus, 4% of males in the Kuwaiti population have G6PD deficiency coexisting with low activity of the UDPGT1 promoter.     

Characterization of a new glucose-6-phosphate dehydrogenase variant: G6PD Central City

Glucose-6-phosphate-dehydrogenase deficiency is the most common disease-producing enzyme deficiency in man. This paper describes a new glucose-6-phosphate-dehydrogenase variant discovered during the evaluation of an episode of acute hemolytic anemia in a 62-year-old black male, which was temporally related to the ingestion of Tolbutamide. The hemolysis resolved within 10 days despite continuation of Tolbutamide. The erythrocyte glucose-6-phosphate-dehydrogenase activity was significantly decreased, and its electrophoretic mobility was indistinguishable from wild type enzyme, though faster on starch gel with tris, borate, and phosphate buffers. The enzyme had a biphasic pH optimum reduced Km for G-6-P and NADP, decreased utilization of deamino-NADP, and reduced Ki for NADPH. Because the kinetic properties of this enzyme were unique, we have designated it as G6PD Central City.     

Glucose-6-phosphate dehydrogenase (G6PD) deficiency in Yogyakarta and its surrounding areas

Glucose-6-phosphate dehydrogenase (G6PD) is one of the enzymes needed by the erythrocyte to generate ATP from ADP. Deficiency of this enzyme can lead to hemolysis of red blood cells. Being a malaria endemic area, Indonesia possibly has a high incidence of G6PD deficiency. It is estimated that 2-6% of the population are carriers. In 1996, we detected 145 neonates with G6PD deficiency using the formazan ring method. Among the males, 6.2% had moderate and 1.4% had low enzyme activity; females had enzyme activity in the normal range. Using the Sigma kit, Tashimi et al in 1995 examined 111 neonates in Yogyakarta, none of which was identified as "deficient". There was no correlation between erythrocyte hemolysis and G6PD enzyme content. Interestingly, using the same Sigma kit. Soro et al in 1994 found that among 134 individuals of Batak descent, 10 males (43.48%) and 9 females (8.11%) were G6PD deficient. These were similar to the results reported by Pramuji et al in 1995 for the people around Palembang. Since the G6PD gene is located on the X chromosome, this is a peculiar result thus further studies need to be done. In cooperation with Harvard University, Sumantri et al in 1995 described 14% as carriers. Molecular analysis among these 16 Javanese males showed the following mutations--nt563 (C->T) in 5 cases, nt1376 (G->T) in 3 cases, nt487 (G->A) in 2 cases, nt1311 (C->T) in 1 case with the remaining variants unknown.     

Molecular heterogeneity of glucose-6-phosphate dehydrogenase (G6PD) variants in the south of Thailand and identification of a novel variant (G6PD Songklanagarind)

Two hundred and twenty-five G6PD-deficient subjects in Songklanagarind Hospital in the south of Thailand comprising 210 males and 15 females were studied. Neonatal jaundice was detected in 85% of these patients. Acute hemolysis related to infection was detected in 17.3% of the G6PD-deficient subjects. Drug-induced acute hemolysis was detected in 1.8% and favism was observed in 3.6% of G6PD-deficient patients. The molecular analysis was performed on 134 G6PD-deficient individuals by a combination of PCR-RFLP, multiplex polymerase chain reaction by multiple tandem forward primers and a common reverse primer assay (MPTP) and DNA sequencing to characterize the mutations of the samples with abnormal MPTP bands. We found 10 different missense G6PD mutations and the three most common variants were G6PD Viangchan 871,G-->A (31.3%), G6PD Kaiping 1388,G-->A (20.1%) and G6PD Mahidol 487,G-->A (17.2%) followed by G6PD Canton 1376,G-->T (9.7%), G6PD Union 1360,C-->T (2.2%), G6PD Gaohe 95,A-->G (1.5%), G6PD Quing Yuan 392,G-->T (0.7%), G6PD Mediterranean 563,C-->T (0.7%), G6PD Songklanagarind 196,T-->A (0.7%), silent mutation 1311,C-->T (6.7%), and uncharacterized variant (9%). A novel missense mutation at codon 196, TTC-->ATC in exon 4 of the G6PD gene predicting a single amino acid substitution, Phe66Ile was identified and we designated this novel class II variant as G6PD Songklanagarind. The G6PD variants among the Thais in the southern part are heterogeneous and G6PD Viangchan, Kaiping, Mahidol, and Canton variants account for about 78% of the cases. Our findings provide some evidence that G6PD Viangchan and Mahidol are common Southeast Asian variants and support the theory of genetic drifts throughout Southeast Asia.     

Glucose-6-phosphate dehydrogenase (G6PD) deficiency in southern Italy: a case of G6PD A(-) associated with favism

During a routine screening for G6PD deficiency in the Province of Matera (Southern Italy), an eleven-year-old boy was brought to our attention who had fever obviously caused by a viral infection, but who also had hepatosplenomegaly and haemoglobinuria. The boy had previously experienced two severe haemolytic attacks. At the age of six months severe haemolysis occurred after the ingestion of cooked fava beans. At the age of seven years, the haemolytic episode was very likely triggered by oral administration of co-trimoxazole. The G6PD activity level in erythrocyte lysate was clearly defective (25% of normal). The electrophoretic mobility of G6PD was 110% of normal. These data together with those obtained from biochemical and molecular characterisation allowed the variant to be identified as G6PD A(-). This is the first report of an association between the African type G6PD deficiency variant and favism.     

Glucose-6-phosphate dehydrogenase (G6PD) deficiency--a cause of anaemia in pregnant women

Glucose-6-phosphate dehydrogenase (G6PD) is one of the most important cytoprotective enzymes for oxidative stress. The WHO classification of G6PD deficiency, based on enzyme activity and clinical significance, distinguishes five variants. Chronic haemolytic process is rare and the main factors causing haemolysis are: infections, substances derived from plants, drugs with high oxidation-reduction potential, stress, ketoacidosis in diabetes and surgery operations. We report two cases of women belonging to the class 3 of the WHO classification in whom haemolysis occured during pregnancy. One of the patients developed two incidents of haemolytic anaemia. The cause of the first episode, nine months before pregnancy, was probably infection of the urinary tract caused by Escherichia coli, but the influence of the drugs also cannot be excluded. Because of the genetic background of this enzymopathy we also examined members of the patients, families but did not find any evidence of G6PD deficiency among them. The reported cases indicate that haemolytic anaemia caused by G6PD deficiency may occur during pregnancy what can lead to many not only haematological but also serious obstetrical complications such as infertility, fetus malformations and even its death. We also draw attention to several difficulties in diagnosing G6PD deficiency especially during haemolysis.     

Molecular characterization of a German variant of glucose-6-phosphate dehydrogenase deficiency (G6PD Aachen)

Glucose-6-phosphate dehydrogenase (G6PD) deficiency is an X-chromosome-linked hereditary disorder. Clinically, patients with G6PD deficiency often present with drug- or food-induced hemolytic crises or neonatal jaundice. G6PD is involved in the generation of NADPH and reduced glutathione. In contrast to American, Mediterranean, and African ancestries, only few variants are known from Middle and Northern Europe. We describe the molecular characterization of a distinct variant from the northwestern area of Germany, G6PD Aachen. The sequence of the G6PD gene from three afflicted males was found to be hemizygous at cDNA residue 1089 for a C-->G mutation with a predicted amino acid change of Asn363Lys. The 1089 C-->G point mutation is unique, but produces the identical amino acid change found in a Mexican variant of G6PD deficiency, G6PD Loma Linda. This G6PD-deficient variant is caused by a 1089 C-->A mutation. The 363-amino-acid replacement is located outside a known mutation cluster region between amino acid residues 380 and 450, but may disrupt or weaken dimer interactions of G6PD enzyme subunits.     

Two new variants of glucose-6-phosphate dehydrogenase associated with hereditary non-spherocytic hemolytic anemia: G6PD Wayne and G6PD Huron

Two new deficient variants of glucose-6-phosphate dehydrogenase (G6PD) causing hereditary nonspherocytic hemolytic anemia (HNSHA) are described. Both of these are unique and they have been named G6PD Wayne and G6PD Huron. Patients with G6PD Wayne underwent splenectomy and no objective improvement was noted. The patients with G6PD Huron were under medical observation for a considerable period of time without the diagnosis of G6PD deficiency being entertained because the family was of Northern European origin. Since sporadic variants of G6PD causing HNSHA show no special racial predilection, the diagnosis of G6PD deficiency should always be considered in patients with this syndrome.     

Glucose-6-phosphate dehydrogenase (G6PD) mutations database: review of the "old" and update of the new mutations

In the present paper we have updated the G6PD mutations database, including all the last discovered G6PD genetic variants. We underline that the last database has been published by Vulliamy et al. [1] who analytically reported 140 G6PD mutations: along with Vulliamy's database, there are two main sites, such as http://202.120.189.88/mutdb/ and www.LOVD.nl/MR, where almost all G6PD mutations can be found. Compared to the previous mutation reports, in our paper we have included for each mutation some additional information, such as: the secondary structure and the enzyme 3D position involving by mutation, the creation or abolition of a restriction site (with the enzyme involved) and the conservation score associated with each amino acid position. The mutations reported in the present tab have been divided according to the gene's region involved (coding and non-coding) and mutations affecting the coding region in: single, multiple (at least with two bases involved) and deletion. We underline that for the listed mutations, reported in italic, literature doesn't provide all the biochemical or bio-molecular information or the research data. Finally, for the "old" mutations, we tried to verify features previously reported and, when subsequently modified, we updated the specific information using the latest literature data.     

Glucose-6-phosphate dehydrogenase (G-6-PD) mutations in Mexico: four new G-6-PD variants

Screening for mutations at the G-6-PD gene by PCR-SSCP combined with restriction enzyme analysis and DNA sequencing was performed in nine G-6-PD deficient individuals with negative results for the presence of the most frequent G-6-PD mutations previously observed in Mexican population. The variants G-6-PD Valladolid(406T), G-6-PD Durham(713G), and G-6-PD Viangchan(871A) and four new G-6-PD mutant alleles were identified. The new mutations are located at cDNA nt 376 A --> T (126 Asn --> Tyr), nt 770 G --> T (257 Arg --> Leu), nt 1094 G --> A (365 Arg --> His), and nt 1285 A --> G (429 Lys --> Glu) and they were named G-6-PD San Luis Potosi, G-6-PD Zacatecas, G-6-PD Veracruz, and G-6-PD Yucatán, respectively. To date, a total of 18 different G-6-PD variants have been observed in Mexico and several of them are common in Africa, South Europe, and Southeast Asia.     

Two new glucose-6 phosphate dehydrogenase (G6PD) variants associated with hemolytic anemia: G6PD Amman-1 and G6PD Amman-2

Two glucose-6-phosphate dehydrogenase (G6PD) variants were investigated. G6PD Amman-1 was partially purified from the red cells of a patient suffering from recurrent jaundice and spontaneous episodic attacks of severe hemolysis in the absence of oxidant drugs, infection, or fava beans. The enzymatic characteristics of G6PD Amman-1 were markedly reduced activity, fast eletrophoretic mobility, slightly increased km for NADP, normal km for G-6-P, normal heat stability, normal utilization of substrate analogues 2-deoxy G-6-P and deamino-NADP, and a monophasic pH curve with a peak at 8.5 to 9.3. The second variant, G6PD Amman-2, was partially purified from the red cells of a patient suffering from recurrent jaundice with episodic mild hemolysis caused by infection or unknown factors. G6PD Amman-2 characteristics were severely reduced activity, slow electrophoretic mobility, normal km for NADP, decreased km for G-6-P, decreased heat stability, increased utilization of substrate analogues, and a monophasic pH curve with a narrow peak at pH 9.5. The red cell level of reduced glutathione was markedly decreased with twofold increase in the activity of glutathione reductase in the patient with G6PD Amman-2.     

G6PD San Francisco: a new variant of glucose-6-phosphate dehydrogenase associated with congenital nonspherocytic hemolytic anemia

Congenital nonspherocytic hemolytic anemia in an adult male of Scandinavian ancestry was associated with virtual absence of G6PD activity in red cells. Characterization of G6PD purified from leukocytes using standard WHO techniques revealed diminished electrophoretic mobility, marked lability on heating at 46 degrees C, normal pH optimum and utilization of alternate substrates (2-deoxy G6P, D-amino NADP), elevated Km NADP, and striking susceptibility to NADPH inhibition. The variant G6PD, which appears to be unique, has been designated G6PD San Francisco. An unusual feature of the variant enzyme, susceptibility to inactivation by brief periods of dialysis, could be prevented by addition of 200 microM NADP to the dialysis solution. In red cells, where G6PD activity was essentially absent, regeneration of reduced glutathione was totally curtailed in vitro, while in leukocytes, where residual G6PD activity was approximately 60% of normal, hexose monophosphate shunt activity, oxygen consumption during phagocytosis, and bacterial killing were unimpaired. Thus, instability of the variant enzyme rather than its unfavorable kinetics appeared to be an important determinant of abnormal cell function.     

Biochemical and molecular characterization of a new sporadic glucose-6-phosphate dehydrogenase variant described in Italy: G6PD Modena

Summary. A new glucose-6-phosphate dehydrogenase variant detected in an Italian man from the Po delata is described and designated as G6PD Modena. Biochemical characterization of the variant enzyme revealed an activity 21% of normal, a slow electrophoretic mobility, increased K_m value for NADP, decreased K_m value for G6P and a complete absence of NADPH inhibition, which could account for the apparently nonhaemolytic feature of this variant. The cloning and sequencing of the G6PD Modena allele showed a GC transition at nucleotide 844 in exon VIII causing a Asp His amino acid substitution. On the basis of biochemical characterization, G6PD Modena is classified as a genuine variant but it has the same mutation as G6PD Seattle-like.