| 11. |
- Guce, Abigail I., et al.
(författare)
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Catalytic Mechanism of Human alpha-Galactosidase
- 2010
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Ingår i: Journal of Biological Chemistry. - : Elsevier BV. - 0021-9258 .- 1083-351X. ; 285:6, s. 3625-3632
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Tidskriftsartikel (refereegranskat)abstract
- The enzyme alpha-galactosidase (alpha-GAL, also known as alpha-GAL A; E.C. 3.2.1.22) is responsible for the breakdown of alpha-galactosides in the lysosome. Defects in human alpha-GAL lead to the development of Fabry disease, a lysosomal storage disorder characterized by the buildup of alpha-galactosylated substrates in the tissues. alpha-GAL is an active target of clinical research: there are currently two treatment options for Fabry disease, recombinant enzyme replacement therapy (approved in the United States in 2003) and pharmacological chaperone therapy (currently in clinical trials). Previously, we have reported the structure of human alpha-GAL, which revealed the overall structure of the enzyme and established the locations of hundreds of mutations that lead to the development of Fabry disease. Here, we describe the catalytic mechanism of the enzyme derived from x-ray crystal structures of each of the four stages of the double displacement reaction mechanism. Use of a difluoro-alpha-galactopyranoside allowed trapping of a covalent intermediate. The ensemble of structures reveals distortion of the ligand into a S-1(3) skew (or twist) boat conformation in the middle of the reaction cycle. The high resolution structures of each step in the catalytic cycle will allow for improved drug design efforts on alpha-GAL and other glycoside hydrolase family 27 enzymes by developing ligands that specifically target different states of the catalytic cycle. Additionally, the structures revealed a second ligand-binding site suitable for targeting by novel pharmacological chaperones.
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| 12. |
- Presneau, Nadege, et al.
(författare)
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Role of the transcription factor T (brachyury) in the pathogenesis of sporadic chordoma: a genetic and functional-based study
- 2011
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Ingår i: Journal of Pathology. - : Wiley. - 0022-3417. ; 223:3, s. 327-335
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Tidskriftsartikel (refereegranskat)abstract
- A variety of analyses, including fluorescence in situ hybridization (FISH), quantitative PCR (qPCR) and array CGH (aCGH), have been performed on a series of chordomas from 181 patients. Twelve of 181 (7%) tumours displayed amplification of the T locus and an additional two cases showed focal amplification; 70/181 (39%) tumours were polysomic for chromosome 6, and 8/181 (4.5%) primary tumours showed a minor allelic gain of T as assessed by FISH. No germline alteration of the T locus was identified in non-neoplastic tissue from 40 patients. Copy number gain of T was seen in a similar percentage of sacrococcygeal, mobile spine and base of skull tumours. Knockdown of T in the cell line, U-CH1, which showed polysomy of chromosome 6 involving 6q27, resulted in a marked decrease in cell proliferation and morphological features consistent with a senescence-like phenotype. The U-CH1 cell line was validated as representing chordoma by the generation of xenografts, which showed typical chordoma morphology and immunohistochemistry in the NOD/SCID/interleukin 2 receptor [IL2r]gamma(null) mouse model. In conclusion, chromosomal aberrations resulting in gain of the T locus are common in sporadic chordomas and expression of this gene is critical for proliferation of chordoma cells in vitro. Copyright (C) 2010 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.
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| 13. |
- Saxena, Richa, et al.
(författare)
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Genetic variation in GIPR influences the glucose and insulin responses to an oral glucose challenge
- 2010
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Ingår i: Nature Genetics. - : Springer Science and Business Media LLC. - 1061-4036 .- 1546-1718. ; 42:2, s. 142-148
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Tidskriftsartikel (refereegranskat)abstract
- Glucose levels 2 h after an oral glucose challenge are a clinical measure of glucose tolerance used in the diagnosis of type 2 diabetes. We report a meta-analysis of nine genome-wide association studies (n = 15,234 nondiabetic individuals) and a follow-up of 29 independent loci (n = 6,958–30,620). We identify variants at the GIPR locus associated with 2-h glucose level (rs10423928, β (s.e.m.) = 0.09 (0.01) mmol/l per A allele, P = 2.0 × 10−15). The GIPR A-allele carriers also showed decreased insulin secretion (n = 22,492; insulinogenic index, P = 1.0 × 10−17; ratio of insulin to glucose area under the curve, P = 1.3 × 10−16) and diminished incretin effect (n = 804; P = 4.3 × 10−4). We also identified variants at ADCY5 (rs2877716, P = 4.2 × 10−16), VPS13C (rs17271305, P = 4.1 × 10−8), GCKR (rs1260326, P = 7.1 × 10−11) and TCF7L2 (rs7903146, P = 4.2 × 10−10) associated with 2-h glucose. Of the three newly implicated loci (GIPR, ADCY5 and VPS13C), only ADCY5 was found to be associated with type 2 diabetes in collaborating studies (n = 35,869 cases, 89,798 controls, OR = 1.12, 95% CI 1.09–1.15, P = 4.8 × 10−18).
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| 14. |
- Solem, C., et al.
(författare)
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Phosphoglycerate Mutase Is a Highly Efficient Enzyme without Flux Control in Lactococcus lactis
- 2010
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Ingår i: Journal of Molecular Microbiology and Biotechnology. - : S. Karger AG. - 1464-1801 .- 1475-3774. ; 18:3, s. 174-180
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Tidskriftsartikel (refereegranskat)abstract
- The glycolytic enzyme phosphoglycerate mutase (PGM), which catalyzes the conversion of 3-phosphoglycerate to 2-phosphoglycerate, was examined in Lactococcus lactis with respect to its function, kinetics and glycolytic flux control. A library of strains with PGM activities ranging between 15-465% of the wild-type level was constructed by replacing the native promoter of pgm with synthetic promoters of varying strengths. The specific growth rate and glucose flux were found to be maximal at the wild-type level at which PGM had no flux control. Low flux control of PGM was found on mixed acid fluxes at highly reduced PGM activities. At the wild-type level PGM operated very far from V-max. Consequently, in a strain with only 15% PGM activity, the catalytic rate of PGM was almost six times higher than in the wildtype. K-m of PGM for 3-phosphoglycerate was 1.0 m M and k(cat) was 3,200 s(-1). The L. lactis PGM was dependent on 2,3-bisphosphoglyceric acid for activity, which showed that the enzyme is of the dPGM type in accordance with its predicted homology to dPGM enzymes from other organisms. In conclusion, PGM from L. lactis is a highly efficient catalyst, which partially explains why this enzyme has limited control in wild-type L. lactis. Copyright (C) 2010 S. Karger AG, Basel
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| 15. |
- Voight, Benjamin F., et al.
(författare)
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Twelve type 2 diabetes susceptibility loci identified through large-scale association analysis
- 2010
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Ingår i: Nature Genetics. - : Springer Science and Business Media LLC. - 1061-4036 .- 1546-1718. ; 42:7, s. 579-589
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Tidskriftsartikel (refereegranskat)abstract
- By combining genome-wide association data from 8,130 individuals with type 2 diabetes (T2D) and 38,987 controls of European descent and following up previously unidentified meta-analysis signals in a further 34,412 cases and 59,925 controls, we identified 12 new T2D association signals with combined P < 5 x 10(-8). These include a second independent signal at the KCNQ1 locus; the first report, to our knowledge, of an X-chromosomal association (near DUSP9); and a further instance of overlap between loci implicated in monogenic and multifactorial forms of diabetes (at HNF1A). The identified loci affect both beta-cell function and insulin action, and, overall, T2D association signals show evidence of enrichment for genes involved in cell cycle regulation. We also show that a high proportion of T2D susceptibility loci harbor independent association signals influencing apparently unrelated complex traits.
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| 16. |
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