کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
10747844 | 1050253 | 2016 | 8 صفحه PDF | دانلود رایگان |
عنوان انگلیسی مقاله ISI
Intracellular co-localization of the Escherichia coli enterobactin biosynthetic enzymes EntA, EntB, and EntE
دانلود مقاله + سفارش ترجمه
دانلود مقاله ISI انگلیسی
رایگان برای ایرانیان
کلمات کلیدی
PVDFChrome azurol SARCPCAS assayBACTHNRPSDHBCASHRPIPTGDTT2,3-dihydroxybenzoic acid - 2،3-دی هیدروکسی بنزوئیک اسیدenterobactin - EnterobactinSDS-polyacrylamide gel electrophoresis - الکتروفورز ژل SDS-polyacrylamide gelSDS-PAGE - الکتروفورز ژل پلی آکریل آمیدisopropyl β-D-1-thiogalactopyranoside - ایزوپروپیل β-D-1-thiogalactopyranosideProtein-protein interactions - تعاملات پروتئین-پروتئینFerric uptake regulator - تنظیم کننده جذب آهنFur - خزbacterial two-hybrid - دوبرابر باکتریاییpolyvinylidene difluoride - دی فلوئورید پلی وینیلیدینdithiothreitol - دیتیوتریتولnon-ribosomal peptide synthesis - سنتز پپتید غیر ریبوزومیSiderophore - سیدرفورHorseradish peroxidase - پراکسیداز هوررادیش
موضوعات مرتبط
علوم زیستی و بیوفناوری
بیوشیمی، ژنتیک و زیست شناسی مولکولی
زیست شیمی
پیش نمایش صفحه اول مقاله
چکیده انگلیسی
Bacteria utilize small-molecule iron chelators called siderophores to support growth in low-iron environments. The Escherichia coli catecholate siderophore enterobactin is synthesized in the cytoplasm upon iron starvation. Seven enzymes are required for enterobactin biosynthesis: EntA-F, H. Given that EntB-EntE and EntA-EntE interactions have been reported, we investigated a possible EntA-EntB-EntE interaction in E. coli cells. We subcloned the E. coli entA and entB genes into bacterial adenylate cylase two-hybrid (BACTH) vectors allowing for co-expression of EntA and EntB with N-terminal fusions to the adenylate cyclase fragments T18 or T25. BACTH constructs were functionally validated using the CAS assay and growth studies. Co-transformants expressing T18/T25-EntA and T25/T18-EntB exhibited positive two-hybrid signals indicative of an intracellular EntA-EntB interaction. To gain further insights into the interaction interface, we performed computational docking in which an experimentally validated EntA-EntE model was docked to the EntB crystal structure. The resulting model of the EntA-EntB-EntE ternary complex predicted that the IC domain of EntB forms direct contacts with both EntA and EntE. BACTH constructs that expressed the isolated EntB IC domain fused to T18/T25 were prepared in order to investigate interactions with T25/T18-EntA and T25/T18-EntE. CAS assays and growth studies demonstrated that T25-IC co-expressed with the EntB ArCP domain could complement the E. coli entBâ phenotype. In agreement with the ternary complex model, BACTH assays demonstrated that the EntB IC domain interacts with both EntA and EntE.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Biochemical and Biophysical Research Communications - Volume 478, Issue 1, 9 September 2016, Pages 25-32
Journal: Biochemical and Biophysical Research Communications - Volume 478, Issue 1, 9 September 2016, Pages 25-32
نویسندگان
Paknoosh Pakarian, Peter D. Pawelek,