Measuring the strength of interaction between the Ebola fusion peptide and lipid rafts: implications for membrane fusion and virus infection.
The Ebola fusion peptide (EBO₁₆) is a hydrophobic domain that belongs to the GP2 membrane fusion protein of the Ebola virus. It adopts a helical structure in the presence of mimetic membranes that is stabilized by the presence of an aromatic-aromatic interaction established by Trp8 and Phe12. In spite of its infectious cycle becoming better understood recently, several steps still remain unclear, a lacuna that makes it difficult to develop strategies to block infection. In order to gain insight into the mechanism of membrane fusion, we probed the structure, function and energetics of EBO₁₆ and its mutant W8A, in the absence or presence of different lipid membranes, including isolated domain-resistant membranes (DRM), a good experimental model for lipid rafts. The depletion of cholesterol from living mammalian cells reduced the ability of EBO₁₆ to induce lipid mixing. On the other hand, EBO₁₆ was structurally sensitive to interaction with lipid rafts (DRMs), but the same was not observed for W8A mutant. In agreement with these data, W8A showed a poor ability to promote membrane aggregation in comparison to EBO₁₆. Single molecule AFM experiments showed a high affinity force pattern for the interaction of EBO₁₆ and DRM, which seems to be a complex energetic event as observed by the calorimetric profile. Our study is the first to show a strong correlation between the initial step of Ebola virus infection and cholesterol, thus providing a rationale for Ebola virus proteins being co-localized with lipid-raft domains. In all, the results show how small fusion peptide sequences have evolved to adopt highly specific and strong interactions with membrane domains. Such features suggest these processes are excellent targets for therapeutic and vaccine approaches to viral diseases.
Centro Nacional de Ressonância Magnética Nuclear Jiri Jonas, Instituto de Bioquímica Médica, Instituto Nacional de Ciências e Tecnologia de Biologia Estrutural e Bioimagem, Universidade Federal do Rio de Janeiro, Rio de Janeiro, Brazil.
Mônica S
MS
Freitas
Cristian
C
Follmer
Lilian T
LT
Costa
Cecília
C
Vilani
M Lucia
ML
Bianconi
Carlos Alberto
CA
Achete
Jerson L
JL
Silva
D002784:Cholesterol / Q000378:metabolism; D029043:Ebolavirus / Q000472:pathogenicity*; D019142:Hemorrhagic Fever, Ebola / Q000209:etiology*; D008561:Membrane Fusion; D021962:Membrane Microdomains / Q000378:metabolism*; D011485:Protein Binding; D014759:Viral Envelope Proteins; D014760:Viral Fusion Proteins / Q000378:metabolism*; D014777:Virus Diseases
D014759:Viral Envelope Proteins; D014760:Viral Fusion Proteins; D002784:Cholesterol
J Biol Chem. 2007 Sep 14;282(37):27306-27314
17545161
Nat Rev Microbiol. 2006 Jan;4(1):67-76
16357862
Virology. 2000 Mar 1;268(1):1-6
10683320
Proc Natl Acad Sci U S A. 1997 Dec 23;94(26):14764-9
9405687
Mol Biol Cell. 1997 Jan;8(1):47-57
9017594
J Virol. 2002 Jan;76(1):406-10
11739705
Proc Natl Acad Sci U S A. 1998 May 12;95(10):5762-7
9576958
J Biol Chem. 2006 Sep 29;281(39):29278-86
16861222
J Gen Physiol. 2008 May;131(5):503-13
18443361
J Virol. 1998 Mar;72(3):1775-81
9499027
J Virol. 1998 Apr;72(4):3155-60
9525641
J Virol. 2006 Mar;80(6):2815-22
16501090
Biochemistry. 2005 Nov 8;44(44):14626-37
16262262
J Virol. 2005 Jan;79(2):918-26
15613320
Mol Biol Cell. 1996 Nov;7(11):1825-34
8930903
J Cell Biol. 2001 Apr 30;153(3):529-41
11331304
J Virol. 2002 May;76(10):5266-70
11967340
J Virol. 1999 Feb;73(2):1419-26
9882347
J Clin Virol. 2001 Oct;22(3):217-27
11564586
J Cell Biol. 1981 Jun;89(3):674-9
6265470
J Virol. 1999 Oct;73(10):8907-12
10482652
Biopolymers. 2002;66(4):249-60
12491538
J Leukoc Biol. 1999 Jul;66(1):50-8
10410989
Cell. 1987 Jun 5;49(5):659-68
3107838
J Biol Chem. 1996 Jul 5;271(27):16026-34
8663188
J Virol. 2003 Dec;77(24):13433-8
14645601
J Virol. 2006 Apr;80(8):4174-8
16571833
Microbes Infect. 2003 Jun;5(7):639-49
12787740
Cell. 1996 May 17;85(4):477-8
8653783
Cell. 2001 Jul 13;106(1):117-26
11461707
FEBS Lett. 2004 Jul 2;569(1-3):261-6
15225645
Nat Rev Mol Cell Biol. 2000 Oct;1(1):31-9
11413487
Eur J Biochem. 1989 Dec 8;186(1-2):17-22
2598927
Future Virol. 2009;4(6):621-635
20198110
FEBS Lett. 2003 Jan 30;535(1-3):23-8
12560072
J Cell Sci. 2001 Nov;114(Pt 22):3957-65
11739628