The trimer-of-hairpins motif in membrane fusion: Visna virus

Malashkevich, V.N.; Singh, Mona; Kim, Peter

doi:10.1073/pnas.151254798

Cited by 45 publications

(38 citation statements)

References 31 publications

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“…Image reconstruction procedures were carried out essentially as described previously (31), using the initial SFV model provided by Dr. Bomu Wu. We used the Iris explorer software (NAG, Inc., Downers Grove, IL), supplemented with custom-made modules 4 for the three-dimensional visualization, along with Pymol TM (DeLano Scientific LLC; available on the World Wide Web).…”

Section: Methodsmentioning

confidence: 99%

“…Opposite to the class I fusion proteins, which are trimers from the start and in which the fusion-related refolding involves formation and backfolding of ␣-helical bundles, the class II fusion proteins elaborate on homotrimer formation to mediate membrane fusion with target membrane, as discussed by Kielian (1,2) and others (3)(4)(5). In the alphaviruses, here represented by Semliki Forest virus (SFV), 3 the fusion proteins are of class II and essentially lack helical motifs.…”

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confidence: 99%

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The Dynamic Envelope of a Fusion Class II Virus

Haag²,

Sjöberg

et al. 2008

Journal of Biological Chemistry

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In alphaviruses, here represented by Semliki Forest virus, infection requires an acid-responsive spike configuration to facilitate membrane fusion. The creation of this relies on the chaperon function of glycoprotein E2 precursor (p62) and its maturation cleavage into the small external E3 and the membrane-anchored E2 glycoproteins. To reveal how the E3 domain of p62 exerts its control of spike functions, we determine the structure of a p62 cleavage-impaired mutant virus particle (SQL) by electron cryomicroscopy. A comparison with the earlier solved wild type virus structure reveals that the E3 domain of p62 SQL forms a bulky side protrusion in the spike head region. This establishes a gripper over part of domain II of the fusion protein, with a cotter-like connection downward to a hydrophobic cluster in its central ␤-sheet. This finding reevaluates the role of the precursor from being only a provider of a shield over the fusion loop to a structural playmate in formation of the fusogenic architecture.When an enveloped virus infects its target cell, the mechanism usually involves a step with hairpin refolding of the viral fusion protein to promote merging of virus and target membranes. Opposite to the class I fusion proteins, which are trimers from the start and in which the fusion-related refolding involves formation and backfolding of ␣-helical bundles, the class II fusion proteins elaborate on homotrimer formation to mediate membrane fusion with target membrane, as discussed by Kielian (1, 2) and others (3-5). In the alphaviruses, here represented by Semliki Forest virus (SFV), 3 the fusion proteins are of class II and essentially lack helical motifs. Homotrimers of the fusion protein E1, formed in relation to the membrane fusion process, are stable associations (6 -13). The strong interaction would be the driving force for completion of fusion, after low pH and membrane contact trigger. However, it would be suicidal for virus transmission if the fusion protein were allowed to create such a configuration prematurely. The SFV assembly pathway handles this problem by providing a chaperon protein, the precursor of glycoprotein E2, in SFV named p62. The p62 precedes the E1 in the proprecursor sequence (p62-6K-E1; see Scheme 1) and waits in the ER to form dimers with the nascent E1, thereby allowing transport to the Golgi compartment and forestalling E1 self-aggregation (10, 15). The E2 itself, if translocated into ER by a cleavable signal sequence, is not sufficient for the purpose. This was shown with an E3 deletion mutant, where the N-terminal E3 domain of the precursor was exchanged for a cleavable artificial signal sequence to preserve the membrane topology of authentic E2. In this E3 deletion mutant, expressed via a recombinant vaccinia virus, the heterodimerization of the spike proteins was abolished, and the E1 was completely retained in the ER (14). In the early Golgi compartment, the p62-E1 heterodimers may form trimers of dimers (16) that in the trans-Golgi undergo furin-dependent maturation cleavage (9,...

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Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

The Dynamic Envelope of a Fusion Class II Virus

Haag²,

Sjöberg

et al. 2008

Journal of Biological Chemistry

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“…7, which is published as supporting information on the PNAS web site, potential interhelical g Arg to eЈ Glu salt bridges are found in many other short (15-50 aa in length) parallel trimeric coiled-coil domains of intracellular, extracellular, transmembrane, viral, and synthetic proteins. Several members of these autonomous coiled coils are well characterized (9,30,31,(33)(34)(35)(36)(37)(38)(39)(40)(41)(42)(43). Based on the alignment, the sequence motif R 1 -h 2 -x 3 -x 4 -h 5 -E 6 can be deduced (R ϭ Arg; E ϭ Glu, L ϭ Leu; h 1 ϭ Ile, Leu, Val, Met; h 2 ϭ Leu, Ile, Val; x ϭ any amino acid residue).…”

Section: The Sequence Motif R-h-x-x-h-e Is Conserved Among Parallel Tmentioning

confidence: 99%

A conserved trimerization motif controls the topology of short coiled coils

Kammerer

Kostrewa

Progias

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

108

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In recent years, short coiled coils have been used for applications ranging from biomaterial to medical sciences. For many of these applications knowledge of the factors that control the topology of the engineered protein systems is essential. Here, we demonstrate that trimerization of short coiled coils is determined by a distinct structural motif that encompasses specific networks of surface salt bridges and optimal hydrophobic packing interactions. The motif is conserved among intracellular, extracellular, viral, and synthetic proteins and defines a universal molecular determinant for trimer formation of short coiled coils. In addition to being of particular interest for the biotechnological production of candidate therapeutic proteins, these findings may be of interest for viral drug development strategies.protein engineering ͉ sequence-to-structure rules ͉ protein-protein interaction ͉ salt bridges ͉ x-ray crystallography T he potential of short ␣-helical coiled coils for protein engineering, biotechnological, biomaterial, basic research, and medical applications has recently been recognized (1-12). The wide range of applications underscores the need for topological control of coiled-coil peptides. Although seemingly simple, coiled coils can form a variety of different assemblies ranging from dimers to pentamers (13,14). Furthermore, coiled coils can form homomers or heteromers with their chains arranged in a parallel or antiparallel fashion. To understand how side-chainside-chain interactions determine a particular structure, it is important to discriminate the factors responsible for specific interhelical interactions from those that are common to all coiled coils such as the heptad repeat.It is generally acknowledged that the detailed packing geometry of hydrophobic core residues correlates with the oligomerization state (15,16). In dimers the side chains of the residues at a and d positions pack in a manner termed parallel and perpendicular, respectively. In the four-stranded state the dimer mode is reversed. Trimeric coiled coils are characterized by an intermediate geometry of these residues, termed acute. Previous studies found that these oligomerization states are determined in particular by the distribution of isoleucine and leucine residues that prefer the parallel and acute, and the acute and perpendicular geometries, respectively (15, 16).Buried polar residues in hydrophobic interfaces also play an important role in determining the number and orientation of strands in coiled coils. Many two-stranded leucine zippers of transcription regulators contain at least one conserved polar residue. Frequently an asparagine or a lysine residue occupies heptad position a toward the center of the sequence, suggesting a common mechanism for determining dimer specificity in these molecules (17). Polar residues like glutamine and threonine at positions a and d, respectively, can favor trimer formation (18,19). Interhelical interactions between side chains of residues at the e and g positions as well as the packi...

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Section: Introductionmentioning

confidence: 99%

“…In these systems, a viral transmembrane surface protein interacts with a receptor in the host cell plasma membrane, leading to docking of the virus particle on the cell surface. Subsequent conformational changes in the interacting proteins finally lead to complete fusion (Dimitrov, 2000;Malashkevich et al, 2001;Mayer, 2001).A small number of eukaryotic genes has been shown by gene disruption to be essential for zygote formation and likely to be required for the late step in fertilization during which the gamete plasma membranes undergo adhesion and fusion. Mouse CD9, a member of the tetraspanin family of proteins, is an egg protein that is essential for fertility.…”

mentioning

confidence: 99%

TheChlamydomonasFus1 Protein Is Present on the Mating TypeplusFusion Organelle and Required for a Critical Membrane Adhesion Event during Fusion withminusGametes

Misamore

Gupta

Snell³

2003

MBoC

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The molecular mechanisms of the defining event in fertilization, gamete fusion, remain poorly understood. The FUS1 gene in the unicellular, biflagellated green alga Chlamydomonas is one of the few sex-specific eukaryotic genes shown by genetic analysis to be essential for gamete fusion during fertilization. In Chlamydomonas, adhesion and fusion of the plasma membranes of activated mtϩ and mtϪ gametes is accomplished via specialized fusion organelles called mating structures. Herein, we identify the endogenous Fus1 protein, test the idea that Fus1 is at the site of fusion, and identify the step in fusion that requires Fus1. Our results show that Fus1 is a ϳ95-kDa protein present on the external surface of both unactivated and activated mtϩ gametes. Bioassays indicate that adhesion between mating type plus and mating type minus fusion organelles requires Fus1 and that Fus1 is functional only after gamete activation. Finally, immunofluorescence demonstrates that the Fus1 protein is present as an apical patch on unactivated gametes and redistributes during gamete activation over the entire surface of the microvillous-like activated plus mating structure, the fertilization tubule. Thus, Fus1 is present on mtϩ gametes at the site of cell-cell fusion and essential for an early step in the fusion process. INTRODUCTIONGamete fusion defines the beginning of a new organism in all sexual species. In most organisms, the events that precede fusion have been well characterized at the cellular level (Snell and White, 1996;Wassarman et al., 2001;Primakoff and Myles, 2002). Initial cell-cell interactions between sperm surface proteins and extracellular matrix molecules on the egg trigger activation of the sperm and exposure of previously cryptic regions of the plasma membrane proposed to be involved in gamete fusion. Once the sperm has made its way to the egg plasma membrane, the membranes of the two gametes interact more intimately, finally bringing about gamete fusion. Several sperm and egg molecules implicated in activation of the sperm have been identified, and we know much about the morphological events that accompany gamete fusion (Yanagimachi, 1994). On the other hand, little is known about the molecular mechanisms that underlie the late steps in fertilization, when the plasma membranes of the interacting gametes adhere and fuse (for review, see Primakoff and Myles, 2002). In the absence of any purely eukaryotic systems in which bona fide fusion proteins have been identified, the best models for fusion of the external plasma membrane of cells have come from studies of fusion of viruses with their eukaryotic target cells (White, 1996;Eckert and Kim, 2001). In these systems, a viral transmembrane surface protein interacts with a receptor in the host cell plasma membrane, leading to docking of the virus particle on the cell surface. Subsequent conformational changes in the interacting proteins finally lead to complete fusion (Dimitrov, 2000;Malashkevich et al., 2001;Mayer, 2001).A small number of eukaryotic genes has been sho...

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The trimer-of-hairpins motif in membrane fusion: Visna virus

Cited by 45 publications

References 31 publications

The Dynamic Envelope of a Fusion Class II Virus

The Dynamic Envelope of a Fusion Class II Virus

A conserved trimerization motif controls the topology of short coiled coils

TheChlamydomonasFus1 Protein Is Present on the Mating TypeplusFusion Organelle and Required for a Critical Membrane Adhesion Event during Fusion withminusGametes

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