Pro-apoptotic bax-alpha1 synthesis and evidence for beta-sheet to alpha-helix conformational change as triggered by negatively charged lipid membranes.
Solid phase synthesis of Bax-alpha1, the 25 amino acids domain (14TSSEQIMKTGALLLQGFIQDRAGRM38) of the pro-apoptotic Bax protein has been accomplished using Fmoc chemistry. A new fast and harmless protocol is described for complete TFA removal from the purified peptide powder leading to a final purity greater than 98% as controlled by 19F-NMR, UV and MALDI-TOF mass spectrometry. Secondary structure was determined in various solution and membrane media using UV Circular Dichroism. In water solution, Bax-alpha1 is present as a mixture of beta-sheet and unstructured (random coil) conformations. A marked change from beta-sheet to alpha-helix secondary structures is observed upon interaction with negatively charged phospholipids vesicles whereas neutral lipid membranes have no significant effect on the aqueous peptide conformation. Results are discussed in terms of Bax binding to mitochondrial membranes.
Sani
Marc-Antoine
MA
UMR 5144 MOBIOS, CNRS-Université Bordeaux 1, IECB, 33607 Pessac Cedex, France.
Loudet
Cécile
C
Gröbner
Gerhard
G
Dufourc
Erick J
EJ
D000180:Acrylic Resins; D000595:Amino Acid Sequence; D051017:Apoptosis Regulatory Proteins / Q000138:chemical synthesis*; D002942:Circular Dichroism; D004134:Dimyristoylphosphatidylcholine; D007202:Indicators and Reagents; D008081:Liposomes; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D010715:Phosphatidylglycerols; D011487:Protein Conformation; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013056:Spectrophotometry, Ultraviolet
D000180:Acrylic Resins; D051017:Apoptosis Regulatory Proteins; D007202:Indicators and Reagents; D008081:Liposomes; D010715:Phosphatidylglycerols; C002773:dimyristoylphosphatidylglycerol; D004134:Dimyristoylphosphatidylcholine