Rat brain hexokinase: the hydrophobic N-terminus of the mitochondrially bound enzyme is inserted in the lipid bilayer.
Mitochondrially bound rat brain hexokinase was labeled with the photoactivatable reagent, 3-(trifluoromethyl)-3-(m-[125I]iodophenyl)diazirine. This highly hydrophobic reagent is strongly partitioned into the hydrophobic environment of the membrane core, and thus selectively labels segments of a protein that penetrate this region of the membrane. Labeling of hexokinase was shown to be restricted to the N-terminal region of the molecule. Approximately 80% of the radiolabel was removed by treatment of the enzyme with chymotrypsin, which preferentially cleaves a hydrophobic 9-residue sequence at the extreme N-terminus of the enzyme, and it is considered likely that the remaining 20% was associated with two additional hydrophobic residues, immediately adjacent to this segment but not susceptible to cleavage by chymotrypsin. Labeling of the enzyme was shown to be dependent on maintenance of the association with the membrane. These results are consistent with a model in which binding of hexokinase involves insertion of an 11-residue hydrophobic N-terminal "tail," possibly existing in alpha-helical secondary structure, into the hydrophobic core of the membrane.
Xie
G C
GC
Department of Biochemistry, Michigan State University, East Lansing 48824.
Wilson
J E
JE
D000345:Affinity Labels; D000818:Animals; D001389:Azirines; D001665:Binding Sites; D001921:Brain / Q000201:enzymology* / Q000378:metabolism; D002918:Chymotrypsin; D006593:Hexokinase / Q000378:metabolism*; D008051:Lipid Bilayers / Q000378:metabolism*; D008565:Membrane Proteins / Q000378:metabolism*; D008928:Mitochondria / Q000201:enzymology; D018272:Porins; D051381:Rats; D012995:Solubility; D014357:Trypsin; D050994:Voltage-Dependent Anion Channels
D000345:Affinity Labels; D001389:Azirines; D008051:Lipid Bilayers; D008565:Membrane Proteins; D018272:Porins; D050994:Voltage-Dependent Anion Channels; C033174:3-(trifluoromethyl)-3-(3-iodophenyl)diazirine; D006593:Hexokinase; D002918:Chymotrypsin; D014357:Trypsin