کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
8299091 | 1537052 | 2008 | 9 صفحه PDF | دانلود رایگان |
عنوان انگلیسی مقاله ISI
Probing the functional tolerance of the b subunit of Escherichia coli ATP synthase for sequence manipulation through a chimera approach
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کلمات کلیدی
DSCFCCPAcmA9-amino-6-chloro-2-methoxyacridine - 9-آمینو-6-کلرو-2-متکی آکرییدینATP synthase - ATP سنتازcircular dichroism - رنگ تابی دورانیB subunit - زیر واحد BChimeric protein - پروتئین کیمیریکDifferential scanning calorimetry - کالریمتری روبشی افتراقیcarbonyl cyanide p-(trifluoromethoxy)phenylhydrazone - کربونیل سیانید p- (trifluoromethoxy) phenylhydrazoneCoiled coil - کویل کویل شده
موضوعات مرتبط
علوم زیستی و بیوفناوری
علوم کشاورزی و بیولوژیک
دانش گیاه شناسی
پیش نمایش صفحه اول مقاله
چکیده انگلیسی
A dimer of 156-residue b subunits forms the peripheral stator stalk of eubacterial ATP synthase. Dimerization is mediated by a sequence with an unusual 11-residue (hendecad) repeat pattern, implying a right-handed coiled coil structure. We investigated the potential for producing functional chimeras in the b subunit of Escherichia coli ATP synthase by replacing parts of its sequence with corresponding regions of the b subunits from other eubacteria, sequences from other polypeptides having similar hendecad patterns, and sequences forming left-handed coiled coils. Replacement of positions 55-110 with corresponding sequences from Bacillus subtilis and Thermotoga maritima b subunits resulted in fully functional chimeras, judged by support of growth on nonfermentable carbon sources. Extension of the T. maritima sequence N-terminally to position 37 or C-terminally to position 124 resulted in slower but significant growth, indicating retention of some capacity for oxidative phosphorylation. Portions of the dimerization domain between 55 and 95 could be functionally replaced by segments from two other proteins having a hendecad pattern, the distantly related E subunit of the Chlamydia pneumoniae V-type ATPase and the unrelated Ag84 protein of Mycobacterium tuberculosis. Extension of such sequences to position 110 resulted in loss of function. None of the chimeras that incorporated the leucine zipper of yeast GCN4, or other left-handed coiled coils, supported oxidative phosphorylation, but substantial ATP-dependent proton pumping was observed in membrane vesicles prepared from cells expressing such chimeras. Characterization of chimeric soluble b polypeptides in vitro showed their retention of a predominantly helical structure. The T. maritima b subunit chimera melted cooperatively with a midpoint more than 20 °C higher than the normal E. coli sequence. The GCN4 construct melted at a similarly high temperature, but with much reduced cooperativity, suggesting a degree of structural disruption. These studies provide insight into the structural and sequential requirements for stator stalk function.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Biochimica et Biophysica Acta (BBA) - Bioenergetics - Volume 1777, Issues 7â8, JulyâAugust 2008, Pages 583-591
Journal: Biochimica et Biophysica Acta (BBA) - Bioenergetics - Volume 1777, Issues 7â8, JulyâAugust 2008, Pages 583-591
نویسندگان
Yumin Bi, Joel C. Watts, Pamela Krauss Bamford, Lee-Ann K. Briere, Stanley D. Dunn,