کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
5506110 | 1400286 | 2017 | 5 صفحه PDF | دانلود رایگان |
عنوان انگلیسی مقاله ISI
Zn2+-dependent autocatalytic activity of the Bordetella pertussis CyaA-hemolysin
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کلمات کلیدی
موضوعات مرتبط
علوم زیستی و بیوفناوری
بیوشیمی، ژنتیک و زیست شناسی مولکولی
زیست شیمی
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چکیده انگلیسی
Proteolytic degradation of the â¼100-kDa isolated RTX (Repeat-in-ToXin) subdomain (CyaA-RTX) of the Bordetella pertussis CyaA-hemolysin (CyaA-Hly) was evidently detected upon solely-prolonged incubation. Here, a truncated CyaA-Hly fragment (CyaA-HP/BI) containing hydrophobic and acylation regions connected with the first RTX block (BI1015-1088) was constructed as a putative precursor for investigating its potential autocatalysis. The 70-kDa His-tagged CyaA-HP/BI fragment which was over-expressed in Escherichia coli as insoluble aggregate was entirely solubilized with 4 M urea. After re-naturation in a Ni2+-NTA affinity column, the purified-refolded CyaA-HP/BI fragment in HEPES buffer (pH 7.4) supplemented with 2 mM CaCl2 was completely degraded upon incubation at 37 °C for 3 h. Addition of 1,10-phenanthrolineâan inhibitor of Zn2+-dependent metalloproteases markedly reduced the extent of degradation for CyaA-HP/BI and CyaA-RTX, but the degradative effect was clearly enhanced by addition of 100 mM ZnCl2. Structural analysis of a plausible CyaA-HP/BI model revealed a potential Zn2+-binding His-Asp cluster located between the acylation region and RTX-BI1015-1088. Moreover, Arg997âone of the identified cleavage sites of the CyaA-RTX fragment was located in close proximity to the Zn2+-binding catalytic site. Overall results demonstrated for the first time that the observed proteolysis of CyaA-HP/BI and CyaA-RTX fragments is conceivably due to their Zn2+-dependent autocatalytic activity.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Biochemical and Biophysical Research Communications - Volume 485, Issue 4, 15 April 2017, Pages 720-724
Journal: Biochemical and Biophysical Research Communications - Volume 485, Issue 4, 15 April 2017, Pages 720-724
نویسندگان
Veerada Raksanoh, Lalida Shank, Panchika Prangkio, Mattayaus Yentongchai, Somsri Sakdee, Chompounoot Imtong, Chanan Angsuthanasombat,