MEMBRANE PROTEIN FOLDING: THE ROLE OF AMINO ACID SEQUENCE IN SPECIFYING THE STABILITY AND CONFORMATION OF TRANSMEMBRANE OLIGOMERS
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Date
2006-08-03T15:29:41Z
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Johns Hopkins University
Abstract
Proteins carry out essential processes of the cell, and to function properly
they must adopt and maintain their native fold. Both thermodynamic and
structural data for soluble protein have provided insight into the forces that drive
folding in an aqueous environment. However, structural and energetic
characterization for membrane proteins has lagged behind that of soluble
proteins. Therefore, membrane protein model systems have been extremely
valuable to understand the interactions that stabilize and specify membrane
protein conformation. Early thermodynamic studies on membrane proteins
demonstrated that interactions between helices promote and specify the native
fold; and a large scale mutagenesis of a transmembrane dimer, Glycophorin A
(GpA), showed that the amino acid sequence on a single face of the helix
conferred stability to the dimer. When the NMR structure was solved, the motif
highlighted by mutagenesis was found at the dimer interface. Therefore,
preferential interactions at the dimer interface may stabilize and specify the
native fold of a membrane protein.
When this thesis study was commenced, interactions in a single sequence
motif were thought to provide the driving force for association in the GpA dimer.
However, in our studies, we have demonstrated that this motif, albeit critical for
strong dimerization, is neither necessary nor sufficient to drive association. This
work highlights that the entire sequence context provides the framework for the
stability and specificity of transmembrane helix-helix interactions. The correlation
of structural models to energetic measurements strongly suggests that local
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packing defects may be responsible for the perturbation upon amino acid
substitution. However, upon experimental examination of structural changes,
packing defects may also initiate global conformation change in the dimer and
therefore these mutations would perturb interactions between the helices and
between the helices and solvating lipids. The information derived from these
studies on a model membrane protein dimer was also applied to transmembrane
segments that are involved in vesicle fusion. The association of these proteins,
syntaxin and synaptobrevin, may be essential to the process of fusion. Both
homo-dimerization and hetero-dimerization of these proteins is weak but is still
modulated by the amino acid sequence. In this case, a dynamic equilibrium for
protein-protein interactions may be critical for biological function. Therefore,
amino acid sequences can encode both stable and transient protein interactions
that are biologically relevant. Thus, when we study membrane protein structure
and interactions, it is necessary to consider the forces that both drive and repel
folding, since it is the equilibrium of these forces that establishes biologically
functional proteins.