Hydrophobic photolabeling identifies BHA2 as the subunit mediating the interaction of bromelain-solubilized influenza virus hemagglutinin with liposomes at low pH.
To investigate the molecular basis of the low-pH-mediated interaction of the bromelain-solubilized ectodomain of influenza virus hemagglutinin (BHA) with membranes, we have photolabeled BHA in the presence of liposomes with the two carbene-generating, membrane-directed reagents 3-(trifluoromethyl)-3-(m-[125I]iodophenyl)diazirine ([125I]TID) and a new analogue of a phospholipid, 1-palmitoyl-2-[11-[4-[3-(trifluoromethyl)diazirinyl]phenyl][2-3H] undecanoyl]-sn-glycero-3-phosphocholine ([3H]-PTPC/11). With the latter reagent, BHA was labeled in a strictly pH-dependent manner, i.e., at pH 5 only, whereas with [125I]TID, labeling was seen also at pH 7. In all experiments, the label was selectively incorporated into the BHA2 polypeptide, demonstrating that the interaction of BHA with membranes is mediated through this subunit, possibly via its hydrophobic N-terminal segment. Similar experiments with a number of other water-soluble proteins (ovalbumin, carbonic anhydrase, alpha-lactalbumin, trypsin, and soybean trypsin inhibitor) indicate that the ability to interact with liposomes at low pH is not a property specific for BHA but is observed with other, perhaps most, proteins.
Harter
C
C
Laboratorium für Biochemie der Eidgenössischen Technischen Hochschule, ETH-Zentrum, Zürich, Switzerland.
Bächi
T
T
Semenza
G
G
Brunner
J
J
D001389:Azirines / Q000138:chemical synthesis*; D001963:Bromelains; D004910:Erythrocyte Membrane / Q000378:metabolism*; D019267:Hemagglutinin Glycoproteins, Influenza Virus; D006389:Hemagglutinins, Viral; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007202:Indicators and Reagents; D008081:Liposomes; D046911:Macromolecular Substances; D009682:Magnetic Resonance Spectroscopy / Q000379:methods; D010713:Phosphatidylcholines / Q000138:chemical synthesis*; D012995:Solubility; D014316:Tritium
D001389:Azirines; D019267:Hemagglutinin Glycoproteins, Influenza Virus; D006389:Hemagglutinins, Viral; D007202:Indicators and Reagents; D008081:Liposomes; D046911:Macromolecular Substances; D010713:Phosphatidylcholines; D014316:Tritium; C033174:3-(trifluoromethyl)-3-(3-iodophenyl)diazirine; C039009:1-palmitoyl-2-(10-(4-((trifluoromethyl)diazirinyl)phenyl)-8-oxadecanoyl)-sn-glycero-3-phosphocholine; D001963:Bromelains