title: Structure and mutagenesis of the parainfluenza virus 5 hemagglutinin-neuraminidase stalk domain reveals a four-helix bundle and the role of the stalk in fusion promotion.
abstract: Paramyxovirus entry into cells requires the fusion protein (F) and a receptor binding protein (hemagglutinin-neuraminidase [HN], H, or G). The multifunctional HN protein of some paramyxoviruses, besides functioning as the receptor (sialic acid) binding protein (hemagglutinin activity) and the receptor-destroying protein (neuraminidase activity), enhances F activity, presumably by lowering the activation energy required for F to mediate fusion of viral and cellular membranes. Before or upon receptor binding by the HN globular head, F is believed to interact with the HN stalk. Unfortunately, until recently none of the receptor binding protein crystal structures have shown electron density for the stalk domain. Parainfluenza virus 5 (PIV5) HN exists as a noncovalent dimer-of-dimers on the surface of cells, linked by a single disulfide bond in the stalk. Here we present the crystal structure of the PIV5-HN stalk domain at a resolution of 2.65 Å, revealing a four-helix bundle (4HB) with an upper (N-terminal) straight region and a lower (C-terminal) supercoiled part. The hydrophobic core residues are a mix of an 11-mer repeat and a 3- to 4-heptad repeat. To functionally characterize the role of the HN stalk in F interactions and fusion, we designed mutants along the PIV5-HN stalk that are N-glycosylated to physically disrupt F-HN interactions. By extensive study of receptor binding, neuraminidase activity, oligomerization, and fusion-promoting functions of the mutant proteins, we found a correlation between the position of the N-glycosylation mutants on the stalk structure and their neuraminidase activities as well as their abilities to promote fusion.
authors: Department of Molecular Biosciences, Northwestern University, 2205 Tech Drive, Evanston, IL 60208-3500, USA.
authors: Sayantan
authors: S
authors: Bose
authors: Brett D
authors: BD
authors: Welch
authors: Christopher A
authors: CA
authors: Kors
authors: Ping
authors: P
authors: Yuan
authors: Theodore S
authors: TS
authors: Jardetzky
authors: Robert A
authors: RA
authors: Lamb
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D002460:Cell Line; D018360:Crystallography, X-Ray; D006031:Glycosylation; D015707:HN Protein / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D050505:Mutant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D010252:Paramyxoviridae / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D011485:Protein Binding; D011487:Protein Conformation; D055503:Protein Multimerization; D053586:Virus Internalization
chemical_list: D015707:HN Protein; D050505:Mutant Proteins