Simple PCR heteroduplex, SSCP mutation screening methods for the detection of novel catalase mutations in Hungarian patients with type 2 diabetes mellitus
-
Márta Vitai
Abstract
Background: The enzyme catalase is the main regulator of hydrogen peroxide metabolism. Deficiency of catalase may cause high concentrations of hydrogen peroxide and increase the risk of the development of pathologies for which oxidative stress is a contributing factor, for example, type 2 diabetes mellitus. Catalase deficiency has been reported to be associated with increased frequency of diabetes mellitus in a cohort of patients in Hungary. In this cohort, the majority of mutations in the catalase gene occur in exon 2.
Methods: Type 2 diabetic patients (n=308) were evaluated for mutations in intron 1 (81bp), exon 2 (172bp) and intron 2 (13bp) of the catalase gene. Screening for mutations utilized PCR single-strand conformational polymorphism (SSCP) and PCR heteroduplex methods. Verification of detected mutations was by nucleotide sequence analysis.
Results: A total of 11 catalase gene mutations were detected in the 308 subjects (3.57%, p<0.001). Five of the 11 were at two previously reported mutation sites: exon 2 (79) G insertion and (138) GA insertion. Six of the 11 were at five previously unreported catalase mutation sites: intron 1 (60) G→T; intron 2 (7) G→A and (5) G→C; exon 2 (96) T→A; and exon 2 (135) T→A. The novel missense mutations on exon 2 (96 and 135) are associated with 59% and 48% decreased catalase activity, respectively; the novel G→C mutation on intron 2 (5) is associated with a 62% decrease in catalase activity. Mutations detected on intron 1 (60) and intron 2 (7) showed no change in catalase activity. The G→C mutation on intron 2 (5) might be a splicing mutation. The two missense mutations on exon 2 (96) and (135) cause substitutions of amino acids 53 (Asp→Glu) and 66 (Glu→Cys) of the catalase protein. These are close to amino acids that are important for the binding of heme to catalase, 44 (Val) and 72–75 (Arg, Val, Val, His). Changes in heme binding may be responsible for the activity losses.
Conclusion: Mutations that cause decreased catalase activity may contribute to susceptibility to inherited type 2 diabetes mellitus. Exon 2 and neighboring introns of the catalase gene may be minor hot spots for type 2 diabetes mellitus susceptibility mutations.
References
1. Berlett BS, Stadtman ER. Protein oxidation in aging, disease, and oxidative stress. J Biol Chem 1997; 272:20313–6.10.1074/jbc.272.33.20313Search in Google Scholar
2. Hunt JV, Wolff SP. Oxidative glycation and free radical production: a causal mechanism for diabetic complications. Free Radical Res Com 1991; 12:115–8.10.3109/10715769109145775Search in Google Scholar
3. Rösen P, Noeroth PP, King G, Möller W, Trischler HJ, Packer L. The role of oxidative stress in the onset and progression of diabetes and its complications: a summary of a Congress Series sponsored by UNESCO-MCBN, the American Diabetes Association and the German Diabetes Society. Diab Metab Res Rev 2001; 17:189–212.10.1002/dmrr.196Search in Google Scholar
4. Evans JL, Goldfine ID, Maddux BA, Grodsky GM. Oxidative stress and stress activated signaling pathways: an unifying hypothesis of the type 2 diabetes. Endocrine Rev 2002; 23:599–622.10.1210/er.2001-0039Search in Google Scholar
5. Peiro C, Lafuente N, Matesanz N, Cercas E, Llergo JL, Vallejo S, et al. High glucose induces cell death of cultured human aortic smooth muscle cells through the formation of hydrogen peroxide. Br J Pharmacol 2001; 133:969–74.10.1038/sj.bjp.0704184Search in Google Scholar
6. Ellis EA, Guberski DL, Somogyi-Mann M, Grant MB. Increased H2O2, vascular endothelial growth factor and receptors in the relation of the BBZ/WOR diabetic rat. Free Radic Biol Med 2000; 28:91–101.10.1016/S0891-5849(99)00216-6Search in Google Scholar
7. Maechler P, Jornot L, Wollheim CB. Hydrogen peroxide alters mitochondrial activation and insulin secretion in pancreatic beta cells. J Biol Chem 1999; 274:27905–13.10.1074/jbc.274.39.27905Search in Google Scholar
8. Hansen LL, Ikeda Y, Olsen GS, Busch AK, Mosthof L. Insulin signalling is inhibited by micromolar concentrations of H2O2. J Biol Chem 1999; 274:25078–84.10.1074/jbc.274.35.25078Search in Google Scholar
9. Masuoka N, Wakimoto M, Ubuka T, Nakano T. Spectrophotometric determination of hydrogen peroxide: catalase activity and rates of hydrogen peroxide removal by erythrocytes. Clin Chim Acta 1996; 254:101–12.10.1016/0009-8981(96)06374-7Search in Google Scholar
10. Gaetani GF, Ferraris AM, Rolfo M, Mangerini R, Arena S, Kirkman HN. Predominant role of catalase in the disposal of hydrogen peroxide within human erythrocytes. Blood 1996; 87:1595–8.10.1182/blood.V87.4.1595.bloodjournal8741595Search in Google Scholar
11. Góth L, Eaton JW. Hereditary catalase deficiencies and increased risk of diabetes. Lancet 2000; 356:1820–1.10.1016/S0140-6736(00)03238-4Search in Google Scholar
12. Góth L, Shemirani A, Kalmár T. A novel catalase mutation (a GA insertion) causes the Hungarian type A of acatalasemia. Blood Cells Mol Dis 2000; 26:151–4.10.1006/bcmd.2000.0288Search in Google Scholar
13. Góth L, Rass P, Madarasi I. A novel catalase mutation detected by PCR-SSCP, nucleotide sequencing and Western blot analyses is responsible for the type C of Hungarian acatalasemia. Electrophoresis 2001; 22:49–51.10.1002/1522-2683(200101)22:1<49::AID-ELPS49>3.0.CO;2-WSearch in Google Scholar
14. Góth L. A novel catalase mutation (a G insertion in exon 2) causes the type B of the Hungarian acatalasemia. Clin Chim Acta 2001; 311:161–3.10.1016/S0009-8981(01)00609-XSearch in Google Scholar
15. Góth L. A new type of inherited catalase deficiencies. Its characterization and comparison to the Japanese and Swiss type of acatalasemia. Blood Cells Mol Dis 2001; 27:512–7.10.1006/bcmd.2001.0415Search in Google Scholar
16. Góth L. Genetic heterogeneity of the 5′ uncoding region of the catalase gene in Hungarian acatalasemic and hypocatalasemic subjects. Clin Chim Acta 1998; 271:173–8.10.1016/S0009-8981(97)00243-XSearch in Google Scholar
17. Góth L, Lenkey A, Bigler NW. Blood catalase deficiency and diabetes in Hungary. Diabetes Care 2001; 10:1139–40.Search in Google Scholar
18. Chance B, Sies H, Boveris A. Hydrogen peroxide metabolism in mammalian organs. Phys Rev 1979; 59:527–605.Search in Google Scholar
19. Góth L. Characterization of acatalasemia detected in two Hungarian sisters. Enzyme 1992; 46:252–8.10.1159/000468797Search in Google Scholar
20. Heales SJ. Catalase deficiencies, diabetes, and mitochondrial function. Lancet 2001; 357:314.10.1016/S0140-6736(05)71763-3Search in Google Scholar
21. Góth L, Vitai M. The effect of hydrogen peroxide promoted by homocysteine and inherited catalase deficiency in human hypocatalasemic patients. Free Radic Biol Med 2003; 35:882–5.10.1016/S0891-5849(03)00435-0Search in Google Scholar
22. Expert Committee on the Diagnosis and Classification of Diabetes Mellitus. Report of the Expert Committee on the Diagnosis and Classification of Diabetes Mellitus. Diabetes Care 1998;21:S5–19.10.2337/diacare.21.1.S5Search in Google Scholar
23. Góth L, Gorzsás A Kalmár T. A simple PCR-heteroduplex screening method for determination of a common mutation of the catalase gene in Hungary. Clin Chem 2000; 46:1199–2000.10.1093/clinchem/46.8.1199Search in Google Scholar
24. Sambrook J, Fritsch EF, Manniatis T. Molecular cloning, 2nd ed. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory Press, 1989:9.14.Search in Google Scholar
25. Kishimoto Y, Murakami Y, Hayashi K, Takahara S, Sugimura T, Sekiya T. Detection of a common mutation of catalase gene in Japanese acatalasemic patients. Hum Genet 1992; 88:487–90.10.1007/BF00219333Search in Google Scholar
26. Vitai M, Góth L. Reference ranges of normal blood catalase activity and levels in familial hypocatalasemia. Clin Chim Acta 1997; 261:35–42.10.1016/S0009-8981(97)06514-5Search in Google Scholar
27. Wen JK, Osumi T, Hashimoto T, Ogata M. Molecular analysis of human acatalasemia: identification of a splicing mutation. J Mol Biol 1990; 211:383–93.10.1016/0022-2836(90)90359-TSearch in Google Scholar
28. Fita I, Rossmann MG. The active center of catalase. J Mol Biol 1985; 185:21–37.10.1016/0022-2836(85)90180-9Search in Google Scholar
29. Murthy MRN, Reid TJ, Sicignano A, Tanaka N, Rossmann MG. Structure of beef liver catalase. J Mol Biol 1981; 152:465–99.10.1016/0022-2836(81)90254-0Search in Google Scholar
30. Putnam CD, Arvai AS, Bourne Y, Tainer JA. Active and inhibited catalase structures: ligand and NADPH binding and catalytic mechanism. J Mol Biol 2000; 296:295–309.10.1006/jmbi.1999.3458Search in Google Scholar PubMed
©2005 by Walter de Gruyter Berlin New York
Articles in the same Issue
- Contents Volume 43, 2005
- Author Index
- Subject Index
- ProteinChips: the essential tools for proteomic biomarker discovery and future clinical diagnostics
- Protein profiling as a diagnostic tool in clinical chemistry: a review
- Protein biochip systems for the clinical laboratory
- Automation of biochip array technology for quality results
- SELDI-TOF-MS proteomics of breast cancer
- Protein microarrays for the diagnosis of allergic diseases: state-of-the-art and future development
- Separation of human serum proteins using the Beckman-Coulter PF2D™ system: analysis of ion exchange-based first dimension chromatography
- Rapid, accurate genotyping of alcohol dehydrogenase-1B and aldehyde dehydrogenase-2 based on the use of denaturing HPLC
- APOA1 polymorphisms are associated with variations in serum triglyceride concentrations in hypercholesterolemic individuals
- Simple PCR heteroduplex, SSCP mutation screening methods for the detection of novel catalase mutations in Hungarian patients with type 2 diabetes mellitus
- Glycogen phosphorylase BB in acute coronary syndromes
- Alteration in serum leptin correlates with alterations in serum N-telopeptide of collagen type I and serum osteocalcin during the progression of osteoporosis in ovariectomized rats
- Glycemic control in diabetes in three Danish counties
- Atorvastatin suppresses homocysteine formation in stimulated human peripheral blood mononuclear cells
- Buprenorphine detection in biological samples
- The effect of thyroid antibody positivity on reference intervals for thyroid stimulating hormone (TSH) and free thyroxine (FT4) in an aged population
- High-affinity antibodies in a new immunoassay for plasma tissue factor: reduction in apparent intra-individual variation
- Physiological matrix metalloproteinase concentrations in serum during childhood and adolescence, using Luminex® Multiplex technology
- Sensitive immunoassays for the autoantibodies reacting against citrullinated carboxy-terminal telopeptides of type I and type II collagens in patients with rheumatoid arthritis
- Acknowledgement
Articles in the same Issue
- Contents Volume 43, 2005
- Author Index
- Subject Index
- ProteinChips: the essential tools for proteomic biomarker discovery and future clinical diagnostics
- Protein profiling as a diagnostic tool in clinical chemistry: a review
- Protein biochip systems for the clinical laboratory
- Automation of biochip array technology for quality results
- SELDI-TOF-MS proteomics of breast cancer
- Protein microarrays for the diagnosis of allergic diseases: state-of-the-art and future development
- Separation of human serum proteins using the Beckman-Coulter PF2D™ system: analysis of ion exchange-based first dimension chromatography
- Rapid, accurate genotyping of alcohol dehydrogenase-1B and aldehyde dehydrogenase-2 based on the use of denaturing HPLC
- APOA1 polymorphisms are associated with variations in serum triglyceride concentrations in hypercholesterolemic individuals
- Simple PCR heteroduplex, SSCP mutation screening methods for the detection of novel catalase mutations in Hungarian patients with type 2 diabetes mellitus
- Glycogen phosphorylase BB in acute coronary syndromes
- Alteration in serum leptin correlates with alterations in serum N-telopeptide of collagen type I and serum osteocalcin during the progression of osteoporosis in ovariectomized rats
- Glycemic control in diabetes in three Danish counties
- Atorvastatin suppresses homocysteine formation in stimulated human peripheral blood mononuclear cells
- Buprenorphine detection in biological samples
- The effect of thyroid antibody positivity on reference intervals for thyroid stimulating hormone (TSH) and free thyroxine (FT4) in an aged population
- High-affinity antibodies in a new immunoassay for plasma tissue factor: reduction in apparent intra-individual variation
- Physiological matrix metalloproteinase concentrations in serum during childhood and adolescence, using Luminex® Multiplex technology
- Sensitive immunoassays for the autoantibodies reacting against citrullinated carboxy-terminal telopeptides of type I and type II collagens in patients with rheumatoid arthritis
- Acknowledgement
