Influence of heme and importance of the N-terminal part of the protein and physical state of model membranes for the apocytochrome c-lipid interaction.
The interaction between cytochrome c and its heme-free precursor apocytochrome c and chemically prepared fragments of these basic proteins with phosphatidylserine containing model membrane systems was studied by differential scanning calorimetry and carboxyfluorescein release experiments. Addition of apocytochrome c and fragments derived from the N-terminus cause a pronounced and linear decrease of the enthalpy (delta H) of the gel to liquid-crystalline phase transition of dielaidoylphosphatidylserine. In contrast, fragments derived from the C-terminus cause a smaller reduction in delta H; a similar trend was observed for the ability of the fragments to cause an increased carboxyfluorescein release from unilamellar vesicles. In addition, the covalent attachment of the heme at cysteine residues 14 and 17 greatly reduced the ability of both the intact protein and the N-terminal fragments to decrease delta H. Using a protein translocation assay based on large unilamellar vesicles containing enclosed trypsin it was found that at gel state temperatures the ability of apocytochrome c to partially translocate the bilayer (reach the opposite membrane/water interface) was greatly reduced. The implications of these findings for the import mechanism of apocytochrome c in mitochondria are shortly indicated.
Zhou
L X
LX
Department of Biochemistry, University of Utrecht, The Netherlands.
Jordi
W
W
De Kruijff
B
B
D000818:Animals; D001059:Apoproteins; D001692:Biological Transport; D002152:Calorimetry, Differential Scanning; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D003574:Cytochrome c Group; D045304:Cytochromes c; D005452:Fluoresceins / Q000494:pharmacology; D005782:Gels; D006418:Heme / Q000502:physiology; D006736:Horses; D008563:Membrane Lipids; D008954:Models, Biological; D010743:Phospholipids; D011485:Protein Binding; D013329:Structure-Activity Relationship; D013816:Thermodynamics
D001059:Apoproteins; D003574:Cytochrome c Group; D005452:Fluoresceins; D005782:Gels; D008563:Membrane Lipids; D010743:Phospholipids; C024098:6-carboxyfluorescein; D006418:Heme; D045304:Cytochromes c