ACERT Highlights: ACERT's Service and Collaborative Projects

National Institute of General Medical Sciences
	National Biomedical Resource for Advanced ESR Spectroscopy

ACERT's Service and Collaborative Projects

Structural Basis for Membrane Anchoring and Fusion Regulation of the Herpes Simplex Virus Fusogen gB from Crystal Structure and DEER Studies

Viral fusogens merge viral and cell membranes during cell penetration. Their ectodomains drive fusion by undergoing largescale refolding, but little is known about the functionally important regions located within or near the membrane. We performed electron spin resonance studies of full-length glycoprotein B (gB), the fusogen from herpes simplex virus, complemented by the crystal structure. The membrane-proximal (MPR), transmembrane (TMD), and cytoplasmic (CTD) domains form a uniquely folded trimeric pedestal beneath the ectodomain, which balances dynamic flexibility with extensive, stabilizing membrane interactions. The postfusion conformation of the ectodomain suggests that the CTD likewise adopted the postfusion form. However, hyperfusogenic mutations, which destabilize the prefusion state of gB, target key interfaces and structural motifs that reinforce the observed CTD structure. Thus, a similar CTD structure must stabilize gB in its prefusion state. Our data suggest a model for how this dynamic, membrane-dependent 'clamp' controls the fusogenic refolding of gB.

Funding: 1R21AI107171, Burroughs Wellcome Fund, Howard Hughes Medical Institute (EEH); 1F32GM115060 (RSC); P41GM103521 (JHF); R01GM123779 (JHF & ERG); in general, P41GM103403, S10RR029205, and DOE DE-AC02-06CH11357.

Publication: Nat. Struct. Mol. Biol. 25, 416-424 (2018); PMCID: PMC5942590.

Interprotomer distances in the isolated CTD. a, Positions of DEER labeling sites and their distances in the crystal structure of gBΔ 71. For clarity, only residues 801-865 of the CTD are shown. b, Experimental time-domain DEER data (left) and reconstructed interprotomer distance distributions (right) for the CTD mutants S803C, E816C, and E830C. Protein in buffer alone is disordered, with broad, widely varying separation between equivalent residues on different protomers. Conversely, distance distributions for samples with a protein: liposome (P/L) ratio of 1:275 coalesce around shorter separation distances, indicating global organization of the CTD.

Rebecca S. Cooper (Department of Molecular Biology and Microbiology, Tufts University school of Medicine, Boston)
Elka R. Georgieva, Peter P. Borbat, Jack H. Freed (ACERT)
and Ekaterina E. Heldwein (Department of Molecular Biology and Microbiology, Tufts University school of Medicine, Boston)

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ACERT is supported by grant 1R24GM146107 from the National Institute of General Medical Sciences (NIGMS), part of the National Institutes of Health.

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