کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
1981567 | 1539419 | 2015 | 9 صفحه PDF | دانلود رایگان |
عنوان انگلیسی مقاله ISI
Structure-based identification of functional residues in the nucleoside-2â²-O-methylase domain of Bluetongue virus VP4 capping enzyme
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کلمات کلیدی
GTaseAdoMetVSVm7GWNVBTV7-methylguanosine - 7-متیل گوانوزینS-adenosyl methionine - S-آدنوزیل متیونینCapping enzyme - بسته شدن آنزیمJEV - جیوMutagenesis - موتاژنزBluetongue virus - ویروس BluetongueVesicular stomatitis virus - ویروس vesicular stomatitisJapanese encephalitis virus - ویروس آنسفالیت ژاپنیWest Nile virus - ویروس نیل غربیpolymerase complex - پیچیده پلیمریزاسیونGuanylyltransferase - گویینیلترانسفراز
موضوعات مرتبط
علوم زیستی و بیوفناوری
بیوشیمی، ژنتیک و زیست شناسی مولکولی
زیست شیمی
پیش نمایش صفحه اول مقاله
چکیده انگلیسی
Bluetongue virus (BTV) encodes a single capping protein, VP4, which catalyzes all reactions required to generate cap1 structures on nascent viral transcripts. Further, structural analysis by X-ray crystallography indicated each catalytic reaction is arranged as a discrete domain, including a nucleoside-2â²-O-methyltransferase (2â²-O MTase). In this study, we have exploited the structural information to identify the residues that are important for the catalytic activity of 2â²-O MTase of VP4 and their influence on BTV replication. The effect of these mutations on GMP binding, guanylyltransferase (GTase) and methylase activities were analysed by a series of in vitro biochemical assays using recombinant mutant proteins; subsequently their effects on virus replication were assessed by introducing the same mutations in replicating viral genome using a reverse genetics system. Our data showed that single substitution mutations in the catalytic tetrad K-D-K-E were sufficient to abolish 2â²-O MTase activity in vitro and to completely abrogate BTV replication in cells; although these mutants retained the upstream GMP binding, GTase and guanine-N7-methyltransferase activities. Mutations of the surrounding substrate-binding pocket (predicted to recruit cap0) had variable effects on in vitro VP4 capping activity. Only triple but not single substitution mutations of these residues in genome resulted in reduced virus replication kinetics. This is the first report investigating the importance of 2â²-O MTase function for any member of the Reoviridae and highlights the significance of K-D-K-E tetrad and surrounding residues for the efficiency of 2â²-O MTase activity and in turn, for virus fitness.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: FEBS Open Bio - Volume 5, 2015, Pages 138-146
Journal: FEBS Open Bio - Volume 5, 2015, Pages 138-146
نویسندگان
Meredith E. Stewart, Polly Roy,