title: pH-triggered conformational switching of the diphtheria toxin T-domain: the roles of N-terminal histidines.
abstract: pH-induced conformational switching is essential for functioning of diphtheria toxin, which undergoes a membrane insertion/translocation transition triggered by endosomal acidification as a key step of cellular entry. In order to establish the sequence of molecular rearrangements and side-chain protonation accompanying the formation of the membrane-competent state of the toxin's translocation (T) domain, we have developed and applied an integrated approach that combines multiple techniques of computational chemistry [e.g., long-microsecond-range, all-atom molecular dynamics (MD) simulations; continuum electrostatics calculations; and thermodynamic integration (TI)] with several experimental techniques of fluorescence spectroscopy. TI calculations indicate that protonation of H257 causes the greatest destabilization of the native structure (6.9 kcal/mol), which is consistent with our early mutagenesis results. Extensive equilibrium MD simulations with a combined length of over 8 μs demonstrate that histidine protonation, while not accompanied by the loss of structural compactness of the T-domain, nevertheless results in substantial molecular rearrangements characterized by the partial loss of secondary structure due to unfolding of helices TH1 and TH2 and the loss of close contact between the C- and N-terminal segments. The structural changes accompanying the formation of the membrane-competent state ensure an easier exposure of the internal hydrophobic hairpin formed by helices TH8 and TH9, in preparation for its subsequent transmembrane insertion.
authors: Chemistry Department, Carnegie Mellon University, Pittsburgh, PA 15213, USA.
authors: Igor V
authors: IV
authors: Kurnikov
authors: Alexander
authors: A
authors: Kyrychenko
authors: Jose C
authors: JC
authors: Flores-Canales
authors: Mykola V
authors: MV
authors: Rodnin
authors: Nikolay
authors: N
authors: Simakov
authors: Mauricio
authors: M
authors: Vargas-Uribe
authors: Yevgen O
authors: YO
authors: Posokhov
authors: Maria
authors: M
authors: Kurnikova
authors: Alexey S
authors: AS
authors: Ladokhin
mesh_terms: D000494:Allosteric Regulation; D004167:Diphtheria Toxin / Q000737:chemistry* / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D008958:Models, Molecular; D056004:Molecular Dynamics Simulation; D011487:Protein Conformation; D017510:Protein Folding / Q000187:drug effects*; D021381:Protein Transport / Q000187:drug effects; D013050:Spectrometry, Fluorescence; D055672:Static Electricity; D013816:Thermodynamics
chemical_list: D004167:Diphtheria Toxin
keywords: FRET; Förster resonance energy transfer; MD; PB; Poisson–Boltzmann; SASA; TI; acid-induced conformational change; fluorescence; histidine protonation; membrane protein insertion; molecular dynamics; solvent-accessible surface area; thermodynamic integration
references: Biochemistry. 2008 Oct 21;47(42):11077-85
references: 18823129
references: Proc Natl Acad Sci U S A. 1997 Apr 15;94(8):3668-72
references: 9108035
references: J Mol Biol. 1999 Jan 29;285(4):1363-9
references: 9917380
references: Proteins. 1991;11(4):281-96
references: 1758883
references: Biochemistry. 2006 Feb 28;45(8):2629-35
references: 16489756
references: Curr Opin Cell Biol. 2006 Dec;18(6):685-9
references: 17046225
references: Curr Opin Struct Biol. 2006 Aug;16(4):525-30
references: 16765587
references: Proc Natl Acad Sci U S A. 1985 Mar;82(6):1692-6
references: 3856850
references: Proc Natl Acad Sci U S A. 1999 Jul 20;96(15):8467-70
references: 10411898
references: Nat Struct Biol. 2003 Jan;10(1):13-8
references: 12459720
references: J Virol. 2009 May;83(9):4670-7
references: 19244325
references: Proteins. 1994 Jun;19(2):141-9
references: 8090708
references: Biochemistry. 2005 Feb 15;44(6):1856-63
references: 15697211
references: Protein Sci. 1994 Sep;3(9):1464-75
references: 7833808
references: Biochemistry. 2009 Aug 18;48(32):7584-94
references: 19588969
references: Proteins. 1993 Mar;15(3):252-65
references: 7681210
references: J Biol Chem. 1998 Sep 4;273(36):22950-6
references: 9722516
references: J Mol Graph. 1996 Feb;14(1):33-8, 27-8
references: 8744570
references: J Gen Physiol. 2000 Apr;115(4):421-34
references: 10736310
references: J Biol Chem. 2011 Jul 22;286(29):25763-9
references: 21592973
references: Annu Rev Cell Dev Biol. 2003;19:45-70
references: 14570563
references: J Chem Phys. 2005 Feb 1;122(5):54101
references: 15740304
references: J Biol Chem. 2007 Aug 17;282(33):24239-45
references: 17584737
references: Nat Rev Mol Cell Biol. 2008 Jan;9(1):47-59
references: 18097445
references: J Gen Physiol. 1998 Sep;112(3):317-24
references: 9725891
references: Biochim Biophys Acta. 2010 Jun;1804(6):1231-64
references: 20117254
references: JAMA. 2001 Feb 28;285(8):1059-70
references: 11209178
references: J Cell Biol. 1984 May;98(5):1757-62
references: 6144684
references: Biophys J. 2011 Nov 16;101(10):L41-3
references: 22098755
references: Biophys J. 2008 Jun;94(11):4348-57
references: 18310255
references: Structure. 2006 Oct;14(10):1481-7
references: 17027497
references: J Cell Biol. 1987 Dec;105(6 Pt 1):2713-21
references: 2447097
references: Biopolymers. 1983 Dec;22(12):2577-637
references: 6667333
references: Nat Struct Biol. 2003 Jan;10(1):2-3
references: 12490883
references: Proteins. 2006 Nov 15;65(3):712-25
references: 16981200
references: EMBO J. 2011 Aug 31;30(17):3481-500
references: 21878991
references: J Mol Biol. 2010 Sep 10;402(1):1-7
references: 20654627
references: Science. 1996 Aug 9;273(5276):810-2
references: 8670424
references: Biochemistry. 2002 Feb 26;41(8):2475-84
references: 11851393