Influenza hemagglutinin: kinetic control of protein function

Baker, David; Agard, David A.

doi:10.1016/s0969-2126(94)00091-3

Cited by 50 publications

(45 citation statements)

References 12 publications

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“…Since both open and closed forms of SAPLIPs contain the same disulfide bonds (15), the primary role of PSI cystines is probably not to maintain the overall fold but perhaps to confer rigidity. Fusion of adjacent membranes consists of the displacement of lipid from its stable, energetically favorable bilayer environment to an aqueous, high energy intermediate state, and such a transition requires enzymatic action (67). Perhaps the energy required to catalyze this event is related to multimer formation (68) and conformation changes (69,70) related to PSI rigidity/stability dependent on disulfide bonds.…”

Section: Discussionmentioning

confidence: 99%

Structure and Mechanism of the Saposin-like Domain of a Plant Aspartic Protease

Bryksa

Bhaumik

Magracheva

et al. 2011

Journal of Biological Chemistry

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Many plant aspartic proteases contain an additional sequence of ϳ100 amino acids termed the plant-specific insert, which is involved in host defense and vacuolar targeting. Similar to all saposin-like proteins, the plant-specific insert functions via protein-membrane interactions; however, the structural basis for such interactions has not been studied, and the nature of plantspecific insert-mediated membrane disruption has not been characterized. In the present study, the crystal structure of the saposin-like domain of potato aspartic protease was resolved at a resolution of 1.9 Å , revealing an open V-shaped configuration similar to the open structure of human saposin C. Notably, vesicle disruption activity followed Michaelis-Menten-like kinetics, a finding not previously reported for saposin-like proteins including plant-specific inserts. Circular dichroism data suggested that secondary structure was pH-dependent in a fashion similar to influenza A hemagglutinin fusion peptide. Membrane effects characterized by atomic force microscopy and light scattering indicated bilayer solubilization as well as fusogenic activity. Taken together, the present study is the first report to elucidate the membrane interaction mechanism of plant saposin-like domains whereby pH-dependent membrane interactions resulted in bilayer fusogenic activity that probably arose from a viral type pH-dependent helix-kink-helix motif at the plant-specific insert N terminus. Aspartic proteases (APs)2 are characterized by a common bilobal tertiary structure containing two catalytic aspartic acid residues (Asp 32 and Asp 215 in pepsin) within an active site cleft(1, 2). They are found in all higher organisms, and their respective roles are well established, although structural and functional characteristics of APs in plants are least understood. Of practical interest among plant APs are their roles in plant pathogen resistance (3) as well as in senescence and postharvest physiology (4, 5). Plant APs share the common AP bilobal structure; however, some contain an additional sequence of ϳ100 residues inserted within the C-terminal primary structure. These additional amino acids unique to plant APs (6 -8) create an extra domain protruding from the canonical AP molecule (9 -11). This structural oddity among APs is called the plantspecific insert (PSI), also known as the plant-specific sequence, which belongs to the saposin-like protein (SAPLIP) family (12, 13). Plant APs are found in either monomeric or heterodimeric forms (9, 14); the latter result from post-translational proteolysis, which includes the removal of part or all of the PSI, whereas the PSI is retained in monomeric plant APs (6,8).In general, members of the SAPLIP family have various physiological functions, all of which entail membrane interaction (14 -16) manifested in three principal ways: membrane binding, membrane perturbation without permeabilization, and membrane permeabilization (15). Examples of SAPLIP functions include roles in exohydrolase degradation of sphingolipids in the...

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Section: Discussionmentioning

confidence: 99%

Structure and Mechanism of the Saposin-like Domain of a Plant Aspartic Protease

Bryksa

Bhaumik

Magracheva

et al. 2011

Journal of Biological Chemistry

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“…In general, spike glycoproteins present on the surfaces of enveloped viruses are locked into a metastable conformation that is usually obtained through proteolytic processing of a precursor(s) (1,25,55). This processing has never been observed for HCV structural proteins.…”

Section: Discussionmentioning

confidence: 99%

Folding of Hepatitis C Virus E1 Glycoprotein in a Cell-Free System

Merola

Brazzoli²,

Cocchiarella³

et al. 2001

J Virol

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The hepatitis C virus (HCV) envelope proteins, E1 and E2, form noncovalent heterodimers and are leading candidate antigens for a vaccine against HCV. Studies in mammalian cell expression systems have focused primarily on E2 and its folding, whereas knowledge of E1 folding remains fragmentary. We used a cell-free in vitro translation system to study E1 folding and asked whether the flanking proteins, Core and E2, influence this process. We translated the polyprotein precursor, in which the Core is N-terminal to E1, and E2 is C-terminal, and found that when the core protein was present, oxidation of E1 was a slow, E2-independent process. The half-time for E1 oxidation was about 5 h in the presence or absence of E2. In contrast with previous reports, analysis of three constructs of different lengths revealed that the E2 glycoprotein undergoes slow oxidation as well. Unfolded or partially folded E1 bound to the endoplasmic reticulum chaperones calnexin and (with lower efficiency) calreticulin, whereas no binding to BiP/GRP78 or GRP94 could be detected. Release from calnexin and calreticulin was used to assess formation of mature E1. When E1 was expressed in the absence of Core and E2, its oxidation was impaired. We conclude that E1 folding is a process that is affected not only by E2, as previously shown, but also by the Core. The folding of viral proteins can thus depend on complex interactions between neighboring proteins within the polyprotein precursor.

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“…How might these transitions take place? Studies on the fusion mechanism of influenza virus suggest a model for the regulation of HA's conversion from the native to the low-pH-induced conformation (5,20). In this model, the conversion of HA would be regulated by kinetic partitioning between its two conformations.…”

Section: Discussionmentioning

confidence: 99%

Formation and Characterization of the Trimeric Form of the Fusion Protein of Semliki Forest Virus

Gibbons

Ahn

Chatterjee

et al. 2000

J Virol

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Enveloped animal viruses infect cells via fusion of the viral membrane with a host cell membrane. Fusion is mediated by a viral envelope glycoprotein, which for a number of enveloped animal viruses rearranges itself during fusion to form a trimeric ␣-helical coiled-coil structure. This conformational change from the metastable, nonfusogenic form of the spike protein to the highly stable form involved in fusion can be induced by physiological activators of virus fusion and also by a variety of destabilizing conditions. The E1 spike protein subunit of Semliki Forest virus (SFV) triggers membrane fusion upon exposure to mildly acidic pH and forms a homotrimer that appears necessary for fusion. We have here demonstrated that formation of the E1 homotrimer was efficiently triggered under low-pH conditions but not by perturbants such as heat or urea, despite their induction of generalized conformational changes in the E1 and E2 subunits and partial exposure of an acid-specific E1 epitope. We used a sensitive fluorescence assay to show that neither heat nor urea treatment triggered SFV-liposome fusion at neutral pH, although either treatment inactivated subsequent low-pH-triggered fusion activity. Once formed, the low-pH-induced E1 homotrimer was very stable and was only dissociated under harsh conditions such as heating in sodium dodecyl sulfate. Taken together, these data, as well as protein structure predictions, suggest a model in which the less stable native E1 subunit specifically responds to low pH to form the more stable E1 homotrimer via conformational changes different from those of the coiled-coil type of fusion proteins.Enveloped viruses must carry out a fusion event between the viral membrane and the host cell membrane to deliver the infectious genome to the cytoplasm. For viruses such as human immunodeficiency virus type 1, fusion occurs at the cell surface in a pH-independent fashion whereas viruses such as Semliki Forest virus (SFV), a member of the alphavirus family, or influenza virus, a myxovirus, fuse via a low-pH-dependent reaction in the endosome. Each of these viruses has integral membrane spike proteins incorporated in the virus envelope that drive membrane fusion in response to the appropriate environmental stimulus, such as receptor binding or a pH change. Proper regulation of the fusogenic properties of these viral spike proteins during virus biosynthesis and maturation is critical to successful virus production and infection of cells. The necessary conformational changes of the proteins that carry out the fusion events have been extensively studied for viruses such as influenza virus and SFV.The influenza virus hemagglutinin (HA) is emblematic of a number of virus fusion proteins with common features. The biosynthetic precursor of HA is a trimer termed HA0 that is activated by proteolytic cleavage to produce a trimer of two disulfide-linked subunits, the receptor-binding HA1 polypeptide and the transmembrane HA2 polypeptide (reviewed in references 26, 61, and 62). Exposure to low pH triggers...

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Influenza hemagglutinin: kinetic control of protein function

Abstract: In response to decreased pH, influenza hemagglutinin changes to a more stable conformation. Such changes, which can be controlled thermodynamically or kinetically, are the method by which many biological 'switches' are thrown.

Cited by 50 publications

References 12 publications

Structure and Mechanism of the Saposin-like Domain of a Plant Aspartic Protease

Structure and Mechanism of the Saposin-like Domain of a Plant Aspartic Protease

Folding of Hepatitis C Virus E1 Glycoprotein in a Cell-Free System

Formation and Characterization of the Trimeric Form of the Fusion Protein of Semliki Forest Virus

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