Secondary Structure and Membrane Topology of the Full‐Length Dengue Virus NS4B in Micelles

Abstract

Dengue virus nonstructural protein 4B (NS4B) is a membrane protein consisting of 248 residues with a crucial role in virus replication and interference with the host innate immunity. The dengue virus serotype 3 NS4B was reconstituted into lyso-myristoyl phosphatidylglycerol (LMPG) micelles. Backbone resonance assignment of NS4B was obtained using conventional solution NMR experiments. Further studies suggested that NS4B contained eleven helices and six of them form five potential transmembrane regions. This study provides atomic level information for an important drug target to control flavivirus infections.

Secondary structure and topology of the transmembrane domain of Syndecan‐2 in detergent micelles

“…SDC2‐TM was purified in the presence of DPC micelles using the method described previously . Briefly, the cells were broken in the buffer with 1% DPC by sonication.…”

“…To understand the dynamics of SDC2-TM in DPC micelles, 15 N T 1 , T 2 , and steady-state heteronuclear NOE (hetNOE) experiments [25] were collected on a 700 MHz magnet using a 15 N-labeled sample with a concentration of 0.3 mM. To obtain T 1 values, one pseudo-3D dataset with several relaxation delays (0.05, 0.1, 0.3, 0.5, 0.8, 1.0, 1.2, 1.6, 2.0, and 2.5 s) was collected and processed.…”

“…1B). Backbone resonances including 15 N, 13 Ca, and 13 C' of residues from syndecan-2 excluding the artificial amino acids at the termini were fully assigned. Most Ha resonances were assigned based on a 3D-HBHACONH experiment.…”

“…2). 15 N-longitudinal relaxation time T 1 , spin-spin relaxation time T 2, and hetNOEs were obtained to understand the flexibility of SCN2-TM in DPC micelles (Fig. 3).…”

“…H-D exchange of SDC2-TM in DPC micelles. (A) 1 H-15 N-HSQC of SDC2-TM in D 2 O at different time were collected and overlaid. The reference spectrum (0 min) used for assignment was collected using a different sample in 10% D 2 O without lyophilization-a step might affect chemical shifts of few residues.…”