Primary structure of peptides and ion channels. Role of amino acid side chains in voltage gating of melittin channels.
Melittin produces a voltage-dependent increase in the conductance of planar lipid bilayers. The conductance increases when the side of the membrane to which melittin has been added (cis-side) is made positive. This paper reports observations on the effect of modifying two positively charged amino acid residues within the NH2-terminal region of the molecule: lysine at position 7 (K7), and the NH2-terminal glycine (G1). We have synthesized melittin analogues in which K7 is replaced by asparagine (K7-N), G1 is blocked by a formyl group (G1-f), and in which both modifications of the parent compound were introduced (G1-f, K7-N). The time required to reach peak conductance during a constant voltage pulse was shorter in membranes exposed to the analogues than in membranes modified by melittin. The apparent number of monomers producing a conducting unit for [K7-N]-melittin and [G1-f]-melittin, eight, was found to be greater than the one for [G1-f], K7-N]-melittin and for melittin itself, four. The apparent gating charge per monomer was less for the analogues, 0.5-0.3 than for melittin, one. Essentially similar results were obtained with melittin analogues in which the charge on K7 or G1 or both was blocked by an uncharged N-linked spin label. These results show that the positive charges in the NH2-terminal region of melittin play a major but not exclusive role in the voltage gating of melittin channels in bilayers.
Department of Cellular Molecular Physiology, Harvard Medical School, Boston, Massachusetts 02115.
M T
MT
Tosteson
O
O
Alvarez
W
W
Hubbell
R M
RM
Bieganski
C
C
Attenbach
L H
LH
Caporales
J J
JJ
Levy
R F
RF
Nutt
M
M
Rosenblatt
D C
DC
Tosteson
D000595:Amino Acid Sequence; D015640:Ion Channel Gating; D007473:Ion Channels / Q000502:physiology*; D007700:Kinetics; D008555:Melitten / Q000138:chemical synthesis / Q000737:chemistry*; D008969:Molecular Sequence Data; D010455:Peptides / Q000138:chemical synthesis; D013329:Structure-Activity Relationship
D007473:Ion Channels; D010455:Peptides; D008555:Melitten