Identification of the critical linker residues conferring differences in the compactness of NS5 from Dengue virus serotype 4 and NS5 from Dengue virus serotypes 1-3

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• 作者：Malathy Sony Subramanian Manimekalai ; Wuan Geok Saw ; Ankita Pan ; Ardina Grü ; ber and Gerhard Grü ; ber
• 关键词：flavivirus ; dengue ; nonstructural proteins ; viral polymerase ; methyltransferase ; small-angle X-ray scattering ; protein flexibility
• 刊名：Acta Crystallographica Section D
• 出版年：2016
• 出版时间：June 2016
• 年：2016
• 卷：72
• 期：6
• 页码：795-807
• 全文大小：2355K
• ISSN：1399-0047

文摘

Dengue virus (DENV) nonstructural protein 5 (NS5) consists of a methyltransferase (MTase) domain and an RNA-dependent RNA polymerase (RdRp) domain. The cross-talk between these domains occurs via a ten-residue linker. Recent solution studies of DENV NS5 from all four serotypes (DENV-1 to DENV-4) showed that NS5 adopts multiple conformations owing to its flexible linker and that DENV-4 NS5 is more compact and less flexible compared with NS5 from DENV-1 to DENV-3 [Saw et al. (2015), Acta Cryst. D71, 2309–2327]. Here, using a variety of single, double, triple and quadruple mutants of DENV-4 NS5 combined with solution X-ray scattering studies, insight into the critical residues responsible for the differential flexibility of DENV-4 NS5 is presented. The DENV-4 NS5 mutants K271T and S266N/T267A as well as the deletion mutant ΔS₂₆₆T₂₆₇ showed enlarged dimensions and flexibility similar to those of DENV-3 NS5. The data indicate that the residues Lys271, Ser266 and Thr267 are important for the compactness of DENV-4 NS5 and therefore may be critical for the regulation of virus replication. Furthermore, quantitative characterization of the flexibility of these DENV-4 NS5 linker mutants using the ensemble-optimization method revealed that these mutants possess a similar conformational distribution to DENV-3 NS5, confirming that these residues in the linker region cause the higher compactness of DENV-4 NS5.