| 1. |
- Endo, Satoshi, et al.
(författare)
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Instability of C154Y variant of aldo-keto reductase 1C3
- 2017
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Ingår i: Chemico-Biological Interactions. - : Elsevier. - 0009-2797 .- 1872-7786. ; 276, s. 194-202
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Tidskriftsartikel (refereegranskat)abstract
- Aldo-keto reductase (AKR) 1C3 is a cytosolic enzyme that metabolizes steroids, prostaglandins, toxic aldehydes and drugs. Recently, some nonsynonymous single nucleotide polymorphisms of AKR1C3 have been suggested to impact steroid and drug metabolism. In this study, we examined the effects of C154Y and L159V variants of AKR1C3 on stability and function of the enzyme. Both variants had been detected in patients with the neurodegenerative disease amyotrophic lateral sclerosis. Recombinant wild-type (WT), C154Y and L159V enzymes were similar in specific activity, but C154Y displayed much lower thermostability than WT and L159V. C154Y was inactivated by 10-min incubation at >25 °C, and about 90% of its activity was lost at 40 °C. Differential scanning fluorimetry revealed that Tm (thermal denaturation midpoint) of C154Y was lower than that of WT. In order to study the cause of thermosensitivity of C154Y, we prepared C154F and C154S mutant AKR1C3s. Like C154Y, C154F was highly sensitive to thermal inactivation, whereas C154S showed almost the same thermostability as WT. The C154F and C154Y variants induced secondary and tertiary structural changes in AKR1C3 at 40 °C as reflected by their altered circular dichroism and 8-anilinonaphthalene-1-sulfonate fluorescence characteristics. These results suggest that the replacement of C154 with a residue possessing a bulky aromatic side-chain impairs the folding of the α-helix containing C154 and its neighboring secondary structures, leading to low thermostability of AKR1C3. AKR1C3 metabolizes cytotoxic 4-oxo-2-nonenal into a less toxic metabolite, and overexpression of WT in HEK293 cells alleviated the 4-oxo-2-nonenal-induced cytotoxicity. In contrast, the overexpression of C154Y in the cells did not show such a significant protective effect, suggesting that C154Y is unstable in cells.
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| 2. |
- Takata, Tohru, et al.
(författare)
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Presence of both heterogeneous vancomycin-intermediate resistance and beta-lactam antibiotic-induced vancomycin resistance phenotypes is associated with the outcome in methicillin-resistant Staphylococcus aureus bloodstream infection
- 2013
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Ingår i: Scandinavian Journal of Infectious Diseases. - : Informa UK Limited. - 0036-5548 .- 1651-1980. ; 45:3, s. 203-212
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Tidskriftsartikel (refereegranskat)abstract
- Background: Although the individual expression of heterogeneous vancomycin-intermediate resistance (hVISA) and beta-lactam antibiotic-induced vancomycin resistance (BIVR) phenotypes has been associated with treatment failure and recurrence in methicillin-resistant Staphylococcus aureus (MRSA) infections, the effect of the co-expression of these phenotypic profiles on clinical outcome has not been fully elucidated. The aim of this study was to determine the impact of the combination of hVISA and BIVR phenotypes on the clinical outcome in MRSA bacteremia. Methods: One hundred and sixty-two MRSA blood isolates from a 21-y period, 1987-2007, were randomly selected. Screening for hVISA was done by the macromethod Etest and confirmed by population analysis profiles. BIVR was identified using Mu3 agar containing 4 mu g/ml of vancomycin. Results: Thirty (18.5%) and 39 (24.1%) of the 162 MRSA blood isolates were positive for the hVISA and BIVR phenotypes, respectively. Eighteen (11.1%) isolates possessed both hVISA and BIVR phenotypes (hVISA(+)/BIVR(+)). In a subset of patients who received initial treatment with glycopeptides, only the patients whose isolates were hVISA(+)/BIVR(+) displayed a significantly higher mortality rate in comparison to those with non-hVISA(+)/BIVR(+) (80.0% vs 31.3%, p = 0.004). The presence of both hVISA and BIVR phenotypes was a predictor of mortality using a logistic regression analysis (p = 0.025). Conclusions: The combined phenotype of hVISA and BIVR was associated with a higher probability of mortality in patients with MRSA bacteremia. Further prospective studies are warranted to delineate the clinical significance of the combined phenotype of hVISA and BIVR.
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