Research Interests
My research interests are focused on two areas of computational study of
protein aggregation and unfolding. The first is focused on the assembly
of Abeta amyloids, which cause Alzheimer's disease. The second involves
the computational investigation of forced (mechanical) unfolding of
proteins. The proposed research program is based on the all-atom
molecular dynamics (MD) simulations of proteins or peptides in explicit
solvent. Both topics are highly
important for understanding the molecular aspects
of Alzheimer's disease and mechanical functions of proteins in living
organisms.
Aggregation of Abeta peptides
The purpose of the computational studies of aggregation
is to understand, at molecular level, the formation of Abeta
amyloid fibrils, which
triggers the onset of Alzheimer's disease. It is recognized that
the development of effective therapeutic treatments demands thorough
understanding of the
kinetics of Abeta peptides' aggregation as well as their fibril
structures.
Recent experiments provided important information on
Abeta fibrils. Solid-state NMR studies [1-3]
showed that Abeta peptides (including the wild-type
Abeta1-40 and the truncated fragment Abeta10-35)
form parallel
in-registry beta-sheets (a two-dimensional structure).
Three-dimensional structural arrangements of
Abeta1-40 and Abeta10-35 fibrils have also been
proposed based on simple energetic considerations [2,4].
In contrast, short
fragments of Abeta peptides (such as Abeta16-22 and
Abeta34-42}) form antiparallel in-registry beta-sheets [5,6].
The kinetics of the assembly of Abeta oligomers (precursors to amyloid
fibrils) has been probed by Teplow and coworkers [7].
They found that in all eighteen
Abeta peptides, which differ in length and residue substitutions, a
transient on-pathway alpha-helical intermediate is observed.
Nevertheless, the molecular basis for the experimental results described
above remains poorly understood. To address these issues we initiated
MD studies of aggregation of fragments of Abeta
peptides ([8,9], see also commentary [10]). For the first time the
formation of Abeta oligomers was observed in
silica, starting with random initial conditions.
In accord with experimental data, MD simulations showed that
antiparallel orientation of Abeta16-22
peptides is energetically preferred.
Dramatic conformational
changes due to interpeptide interactions were observed, which include
obligatory alpha-helical intermediate.
We established that oligomerization of Abeta peptides
is initially driven by hydrophobic interactions. However,
electrostatic interactions confer antiparallel
registry of peptides, which results in further decrease in potential
energy. This
kinetic mechanism of Abeta oligomerization may
represent a universal pathway of amyloid fibril formation.
Force-induced unfolding of proteins
Force-induced (mechanical) unfolding of proteins is implicated in muscle
contraction and
relaxation [11],
cell adhesion and receptor binding [12],
tissue elasticity [11] etc.
For example, titin molecule provides passive elasticity
for muscle tissues and is composed of about 300 folded domains, among
which are immunoglobulin-type (Ig) and fibronectin-type (Fn) domains.
Modular construction transforms titin into
a complex nonlinear spring [13].
Another mechanically active protein, fibronectin, is a part of
extracellular matrix (ECM), which determines
elasticity and tensile strength of tissues [11].
Similar to titin, fibronectin includes
several types of domains, some of which (Fn3 domains) incorporate
binding sites for integrin and heparin molecules [14-16].
It is likely that stretching of Fn3 domains changes the affinity
of binding regions and also exposes new "cryptic" association sites
[17-18]. Therefore, mechanical unfolding of
fibronectin controls the assembly of ECM amyloid fibrils, cell adhesion
and migration [12].
Recent atomic force microscopy (AFM) experiments [19] measured, at
single molecule level, the
unfolding forces of various domains in mechanically active proteins,
including titin and fibronectin. The AFM experiments revealed
hierarchic (``one by one'') unfolding of domains in the proteins,
which starts with the weakest one. Interestingly, the stability of protein
domains against thermal or chemical denaturation does not correlate
with the mechanical stability [20].
MD simulations
provided detailed information on the structural events occurring upon
force-induced unfolding of individual domains [21].
Computer simulations performed by us using
low-resolution protein models identified the native
topology as the key factor determining the unfolding pathways and
rapture forces in proteins [22].
In addition, we developed
a novel algorithm, which provides accurate prediction of mechanical
stability and
force-induced unfolding pathways of wild-type proteins [22].
-
Antzutkin, O.N., Balbach, J.J., Leapman, R.D., Rizzo, N.W.,
Reed, J., and Tycko, R. (2000)
Multiple quantum solid-state NMR indicates a parallel,
not antiparallel, organization of beta-sheets in Alzheimer's
beta-amyloid fibrils. Proc. Natl. Acad. Sci. USA 97, 13045-13050.
- Petkova, A.T., Ishii, Y., Balbach, J.J.,
Antzutkin, O.N., Leapman, R.D., Delaglio, F., and Tycko, R.
(2002) A structural model for Alzheimer's beta-amyloid
fibrils based on experimental constraints from solid state NMR.
Proc. Natl. Acad. Sci. USA 99, 16742-16747.
- Burkoth, T.S., Benzinger, T.L.S., Urban, V.,
Morgan, D.M., Gregory, D.M., Thiyagarajan, P., Botto, R.E., Meredith, S.C.,
and Lynn, D.G. (2000) Structure of the beta-Amyloid(10-35) fibril.
J. Am. Chem. Soc. 122, 7883-7889.
- Ma, B. and Nussinov, R. (2002)
Stabilities and conformations of Alzheimer's beta-amyloid
peptide oligomers (Abeta16-22, Abeta16-35,
Abeta10-35): Sequence effects.
Proc. Natl. Acad. Sci. USA 99, 14126-14131.
- Balbach, J.J., Ishii, Y., Antzutkin, O.N.,
Leapman, R.D., Rizzo, N.W., Dyda, F., Reed, J., and Tycko, R. (2000)
Amyloid fibril formation by Abeta16-22, a seven-residue
fragment of the
Alzheimer's beta-amyloid peptide, and structural characterization by
solid state NMR. Biochem. 39, 13748-13759.
-
Lansbury, P.T., Costa, P.R., Griffiths, J.M., Simon, E.J.,
Auger, M., Halverson, K.J., Kocisko, D.A., Hendsch, Z.S.,
Ashburn, T.T., Spencer, R.G.S. et al. (1995) Structural
model for the beta-amyloid fibril based on interstrand alignment
of an antiparallel-sheet comprising a C-terminal peptide.
Nat. Struct. Biol. 2, 990-998.
- Kirkitadze, M.D., Condron, M.M., and Teplow, D.B. (2001)
Identification and characterization of key kinetic
intermediates in amyloid beta-protein fibrillogenesis.
J. Mol. Biol. 312, 1103-1119.
- Klimov, D.K. and Thirumalai, D. (2003)
Dissecting the assembly of Abeta16-22 amyloid
peptides into antiparallel beta-sheets. Structure 11, 295-307.
- Lynn, D.G., Snyder, J.P., Lakdawala, A.S.,
Dong, J., and Lu, K. (2003) Conformational evolution: The wiggling of
peptides into amyloid. Structure 11, 242.
- Balbirnie, M., Grothe, R., and Eisenberg, D.S.
(2001) An amyloid-forming peptide from the yeast prion Sup35
reveals a dehydrated beta-sheet structure for amyloid. Proc. Natl.
Acad. Sci. USA 98, 2375-2380.
-
Massi, F., Klimov, D., Thirumalai, D., and Straub, J. E. (2002)
Charged states rather than propensity for beta-structure
determine enhanced fibrillogenesis in wild-type Alzheimer's
beta-amyloid peptide compared to E22Q Dutch mutant.
Prot. Sci. 11, 1639-1647.
- Erickson, H.P. (1994)
Reversible unfolding of fibronectin type III and immunoglobulin
domains provides the structural basis for stretch and elasticity
of titin and fibronectin. Proc. Natl. Acad. Sci. USA
91, 10114-10118.
- Hynes, R.O. (1999)
The dynamic dialogue between cells and matrices: Implications of
fibronectin's elasticity. Proc. Natl. Acad. Sci. USA 96,
2588-2590.
- Erickson, H.P. (1997)
Stretching single protein modules: Titin is a weird spring. Science
276, 1090-1093.
-
Leahy, D.J., Aukhil, I.L., and Erickson, H.P. (1996)
2.0\AA \ \ crystal structure of a four-domain segment of human
fibronectin encompassing the RGD loop and synergy region. Cell
84, 155-164.
-
Johanssen, S., Svineng, G., Wennerberg, K., Armulik, A., and
Lohikangas, L. (1997) Fibronectin-integrin interactions.
Frontiers Biosci. 2, 126-146.
-
Sharma, A., Askari, J.A., Humphries, M.J., Jones, E.Y., Stuart, D.I. (1999)
Crystal structure of a heparin- and integrin-binding segments of
human fibronectin. Eur. Mol. Biol. Org. J. 18, 1468-1479.
-
Ingham, K.C., Brew, S.A., Huff, S., and Litvinovich, S.V. (1997)
Cryptic self-association sites in type III modules of fibronectin.
J. Biol. Chem. 272, 1718-1724.
-
Litvinovich, S.V., Brew, S.A., Aota, S., Akiyama, S.K.,
Haudenschild, C., and Ingham, K.C. (1998)
Formation of amyloid-like fibrils by self-association of a
partially unfolded fibronectin type {III} module.
J. Mol. Biol. 280, 245-258.
-
Fisher, T.E., Oberhauser, A.F., Carrion-Vazquez, M.,
Marszalek, P.E., and Fernandez, J.M. (1999)
The study of protein mechanics with the atomic force microscope.
Trends in Biochem. Sci. 24, 379-384.
-
Li, H., Oberhouser, A.F., Fowler, S.B., Clarke, J., Fernandez, J.M. (2000)
Atomic force microscopy reveals the mechanical design of a modular
protein. Proc. Natl. Acad. Sci. USA 97, 6527-6531.
-
Isralewitz, B., Gao, M., and Schulten, K. (2001)
Steered molecular dynamics and mechanical functions of proteins.
Curr. Opin. Struct. Biol. 11, 224-230.
-
Klimov, D.K. and Thirumalai, D. (2000)
Native topology determines force-induced unfolding pathways in
globular proteins. Proc. Natl. Acad. Sci. USA 97, 7254-7259.
-
Oberhauser, A.F., Badilla-Fernandez, C.,
Carrion-Vazquez, M., and Fernandez, J.M. (2002)
The mechanical hierarchies of fibronectin observed with
single-molecule AFM. J. Mol. Biol. 319, 433-447.
- Paci, E. and Karplus, M. (1999)
Forced unfolding of fibronectin type 3 modules: An analysis
by biased molecular dynamics simulations. J. Mol. Biol 288,
441-459.
-
Litvinovich, S.V. and Ingham, K.C. (1995)
Interactions between type III domains in the 110 kDa
cell-binding fragment of fibronectin. J. Mol. Biol.
248, 611-626.
- Betts, S. & King, J. (1999) There's a right way and a
wrong way: in vivo and in vitro folding, misfolding and
subunit assembly of the P22 tailspike. Structure 7,
R131-R139.