- Tytuł:
- Spectrophotometric method for the determination of renal ouabain-sensitive H+,K+ -ATPase activity.
- Autorzy:
-
Bełtowski, Jerzy
Wójcicka, Grażyna - Powiązania:
- https://bibliotekanauki.pl/articles/1043791.pdf
- Data publikacji:
- 2002
- Wydawca:
- Polskie Towarzystwo Biochemiczne
- Tematy:
-
leptin
kidney
H+,K+-ATPase
ouabain
Sch 28080
Na+,K+-ATPase - Opis:
- The aim of this work was to develop a method for renal H+,K+-ATPase measurement based on the previously used Na+,K+-ATPase assay (Bełtowski et al.: J Physiol Pharmacol.; 1998, 49: 625-37). ATPase activity was assessed by measuring the amount of inorganic phosphate liberated from ATP by isolated microsomal fraction. Both ouabain-sensitive and ouabain-resistant K+-stimulated and Na+-independent ATPase activity was detected in the renal cortex and medulla. These activities were blocked by 0.2 mM imidazolpyridine derivative, Sch 28080. The method for ouabain- sensitive H+,K+-ATPase assay is characterized by good reproducibility, linearity and recovery. In contrast, the assay for ouabain-resistant H+,K+-ATPase was unsatisfactory, probably due to low activity of this enzyme. Ouabain-sensitive H+,K+-ATPase was stimulated by K+ with Km of 0.26 ± 0.04 mM and 0.69 ± 0.11 mM in cortex and medulla, respectively, and was inhibited by ouabain (Ki of 2.9 ± 0.3 μM in the renal cortex and 1.9 ± 0.4 μM in the renal medulla) and by Sch 28080 (Ki of 1.8 ± 0.5 μM and 2.5 ± 0.9 μM in cortex and medulla, respectively). We found that ouabain-sensitive H+,K+-ATPase accounted for about 12% of total ouabain-sensitive activity in the Na+,K+-ATPase assay. Therefore, we suggest to use Sch 28080 during Na+,K+-ATPase measurement to block H+,K+-ATPase and improve the assay specificity. Leptin administered intraperitoneally (1 mg/kg) decreased renal medullary Na+,K+-ATPase activity by 32.1% at 1 h after injection but had no effect on H+,K+-ATPase activity suggesting that the two renal ouabain-sensitive ATPases are separately regulated.
- Źródło:
-
Acta Biochimica Polonica; 2002, 49, 2; 515-527
0001-527X - Pojawia się w:
- Acta Biochimica Polonica
- Dostawca treści:
- Biblioteka Nauki
Artykuł