TY - JOUR
T1 - The measles virus hemagglutinin stalk
T2 - Structures and functions of the central fusion activation and membrane-proximal segments
AU - Navaratnarajah, Chanakha K.
AU - Kumar, Swati
AU - Generous, Alex
AU - Apte-Sengupta, Swapna
AU - Mateo, Mathieu
AU - Cattaneo, Roberto
PY - 2014/6
Y1 - 2014/6
N2 - The measles virus (MeV) membrane fusion apparatus consists of a fusion protein trimer and an attachment protein tetramer. To trigger membrane fusion, the heads of the MeV attachment protein, hemagglutinin (H), bind cellular receptors while the 96- residue-long H stalk transmits the triggering signal. Structural and functional studies of the triggering mechanism of other paramyxoviruses suggest that receptor binding to their hemagglutinin-neuraminidase (HN) results in signal transmission through the central segments of their stalks. To gain insight into H-stalk structure and function, we individually replaced its residues with cysteine. We then assessed how stable the mutant proteins are, how efficiently they can be cross-linked by disulfide bonds, whether cross-linking results in loss of function, and, in this case, whether disulfide bond reduction restores function. While many residues in the central segment of the stalk and in the spacer segment above it can be efficiently cross-linked by engineered disulfide bonds, we report here that residues 59 to 79 cannot, suggesting that the 20 membrane-proximal residues are not engaged in a tetrameric structure. Rescue-of-function studies by disulfide bond reduction resulted in the redefinition and extension of the central fusion-activation segment as covering residues 84 to 117. In particular, we identified four residues located between positions 92 and 99, the function of which cannot be restored by disulfide bond reduction after cysteine mutagenesis. These mutant H proteins reached the cell surface as complex oligomers but could not trigger membrane fusion. We discuss these observations in the context of the stalk exposure model of membrane fusion triggering by paramyxoviruses.
AB - The measles virus (MeV) membrane fusion apparatus consists of a fusion protein trimer and an attachment protein tetramer. To trigger membrane fusion, the heads of the MeV attachment protein, hemagglutinin (H), bind cellular receptors while the 96- residue-long H stalk transmits the triggering signal. Structural and functional studies of the triggering mechanism of other paramyxoviruses suggest that receptor binding to their hemagglutinin-neuraminidase (HN) results in signal transmission through the central segments of their stalks. To gain insight into H-stalk structure and function, we individually replaced its residues with cysteine. We then assessed how stable the mutant proteins are, how efficiently they can be cross-linked by disulfide bonds, whether cross-linking results in loss of function, and, in this case, whether disulfide bond reduction restores function. While many residues in the central segment of the stalk and in the spacer segment above it can be efficiently cross-linked by engineered disulfide bonds, we report here that residues 59 to 79 cannot, suggesting that the 20 membrane-proximal residues are not engaged in a tetrameric structure. Rescue-of-function studies by disulfide bond reduction resulted in the redefinition and extension of the central fusion-activation segment as covering residues 84 to 117. In particular, we identified four residues located between positions 92 and 99, the function of which cannot be restored by disulfide bond reduction after cysteine mutagenesis. These mutant H proteins reached the cell surface as complex oligomers but could not trigger membrane fusion. We discuss these observations in the context of the stalk exposure model of membrane fusion triggering by paramyxoviruses.
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U2 - 10.1128/JVI.02846-13
DO - 10.1128/JVI.02846-13
M3 - Article
C2 - 24648460
AN - SCOPUS:84899850258
SN - 0022-538X
VL - 88
SP - 6158
EP - 6167
JO - Journal of virology
JF - Journal of virology
IS - 11
ER -