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Biochemical comparison of Anopheles gambiae and human NADPH P450 reductases reveals different 2'-5'-ADP and FMN binding traits.

التفاصيل البيبلوغرافية
العنوان: Biochemical comparison of Anopheles gambiae and human NADPH P450 reductases reveals different 2'-5'-ADP and FMN binding traits.
المؤلفون: Lu-Yun Lian, Philip Widdowson, Lesley A McLaughlin, Mark J I Paine
المصدر: PLoS ONE, Vol 6, Iss 5, p e20574 (2011)
بيانات النشر: Public Library of Science (PLoS), 2011.
سنة النشر: 2011
المجموعة: LCC:Medicine
LCC:Science
مصطلحات موضوعية: Medicine, Science
الوصف: NADPH-cytochrome P450 oxidoreductase (CPR) plays a central role in chemical detoxification and insecticide resistance in Anopheles gambiae, the major vector for malaria. Anopheles gambiae CPR (AgCPR) was initially expressed in Eschericia coli but failed to bind 2',5'-ADP Sepharose. To investigate this unusual trait, we expressed and purified a truncated histidine-tagged version for side-by-side comparisons with human CPR. Close functional similarities were found with respect to the steady state kinetics of cytochrome c reduction, with rates (k(cat)) of 105 s(-1) and 88 s(-1), respectively, for mosquito and human CPR. However, the inhibitory effects of 2',5'-ADP on activity were different; the IC(50) value of AgCPR for 2',5'-ADP was significantly higher (6-10 fold) than human CPR (hCPR) in both phosphate and phosphate-free buffer, indicative of a decrease in affinity for 2',5'-ADP. This was confirmed by isothermal titration calorimetry where binding of 2',5'-ADP to AgCPR (K(d) = 410±18 nM) was ∼10 fold weaker than human CPR (K(d) = 38 nM). Characterisation of the individual AgFMN binding domain revealed much weaker binding of FMN (K(d) = 83±2.0 nM) than the equivalent human domain (K(d) = 23±0.9 nM). Furthermore, AgCPR was an order of magnitude more sensitive than hCPR to the reductase inhibitor diphenyliodonium chloride (IC(50) = 28 µM±2 and 361±31 µM respectively). Taken together, these results reveal unusual biochemical differences between mosquito CPR and the human form in the binding of small molecules that may aid the development of 'smart' insecticides and synergists that selectively target mosquito CPR.
نوع الوثيقة: article
وصف الملف: electronic resource
اللغة: English
تدمد: 1932-6203
Relation: http://europepmc.org/articles/PMC3105092?pdf=render; https://doaj.org/toc/1932-6203
DOI: 10.1371/journal.pone.0020574
URL الوصول: https://doaj.org/article/c078b1c8cc5248999abac58d0c0db96f
رقم الأكسشن: edsdoj.078b1c8cc5248999abac58d0c0db96f
قاعدة البيانات: Directory of Open Access Journals
الوصف
تدمد:19326203
DOI:10.1371/journal.pone.0020574