Nanomedicine

Center for Protein Folding Machinery

2007 Progress Report – Executive Summary

A hallmark of biological nanomachines is their complex and precise architectural design and their dynamic transitions between multiple conformational states inside the cell while performing their functional tasks concurrent with regulation by other cellular factors. A major challenge in characterizing these nanomachines is the lack of established approaches to generate a quantitative description of their conformational adaptability while in action and prediction of their conformational variations in the cellular environment. We believe that such information is critical to performing the rational design of nanomachines with new functionalities.

The specific nanomachine that our NDC focuses on is the type II molecular chaperonin including the eukaryotic chaperonin TRiC/CCT and the archeal chaperonin Mm-cpn. They differ in complexity: while TRiC is made up of 8 distinct subunits in a double-ring arrangement, Mmcpn is made up of 16 identical subunits in two rings. Both share a ring-shaped molecular architecture and both function as nanocontainers that enclose unfolded polypeptides within the central cavity and promote their folding in an ATP-dependent manner. Describing the conformational dynamics of this process is of central importance to understanding and eventually controlling protein folding in the cell. The ultimate goal of our NDC is to be able to engineer chaperonins that will fold any protein of interest and to design molecular adaptors that will facilitate interactions between chaperonins and substrate of interest resulting in proper folding.

In the past year, we have made dramatic progress in the development of novel biophysical and computational methodologies to sample the conformational space of the chaperonins and their complexes with substrates. All the groups in our NDC have been working closely and collaboratively towards this goal. Our efforts place particular emphasis on achieving the integration of biological, physical and computational approaches as represented by our complementary expertise. Our mechanisms of interaction include frequent cross-lab experiments and discussion sessions, exchange visits, Web-based seminars and sub-group meetings that have been instrumental in inter-disciplinary collaboration, training and seeding new ideas for both technological and conceptual advances. Our findings this year have produced several exciting technological breakthroughs. We also discovered previously unknown biological properties of TRiC that have a clear relevance for the Pathways to Medicine projects.

The highlights of our advances are summarized as followings:

• Finding the molecular switch to open and close the TRiC chamber. (Chiu, Ludtke, Sali, Levitt, Frydman)
• Proposing a Two-Stroke allosteric model for the Mm-cpn built-in lid (Frydman, Chiu)
• Detecting the chemical behavior of ATP bound to chaperonins (Moerner, Frydman)
• Determining the specificity of TRiC binding to medically relevant substrates (Frydman)
• Discovering a new type of chaperonin-substrate interaction (Moerner, Frydman)
• Modeling the effect of the chaperonin cavity on folding of bound proteins (Pande, Frydman)
• Progressing towards the design of a molecular adaptor between chaperonins and substrate interfaces (Kortemme, Frydman)
• New Evidence shows chaperonins as misfolding and aggregation inhibitor for Huntington Disease (Frydman)
• New Evidence shows binding of archeal chaperonins to lens protein (King, Frydman)
• A Novel image reconstruction method reveals a conformational mixture during a folding reaction inside the chaperonin (Ludtke)
• New computational method for predicting sequential ordering and molecular interactions among subunits in TRiC (Sali, Frydman)
• Integrating small angle X-ray scattering data and modeling for structure determination of nanomachine and substrates (Sali, Adams)
• Developing new tools to model complex protein interaction interfaces (Kortemme)
• Progress in X-ray diffraction of the Mm-cpn chaperonin and the TRiC domains (Adams, Frydman)
• Model the dynamic behavior of nanomachines (Pande)
• Producing animations of protein folding event (Gossard, Chiu)

In the next year, we plan to continue with the highly successful lines of research we have already established. Our basic strategy is to focus on the four major research thrusts: quantitative “basic science” to define the chaperonin nanomachine properties through both experiments and simulations; engineering and design to manipulate the interactions between chaperonins and substrate of interest; concerted engagement in the pathway to medicine; technology innovation to push the envelope in the study and manipulation of biological nanomachines. We feel these provide a powerful combination that will eventually allow for innovative new therapeutic and research avenues. In addition, we fully recognize the freedom of this NDC to explore new avenues and high risk type of research. Using Supplementary Funds, we will explore further medical applications with some of these new findings.

Current year publications

Spiess, C; Miller, E.J. McClellan, A.J. and J. Frydman (2006). Identification of the TRiC/CCT substrate binding sites uncovers the function of subunit diversity in eukaryotic chaperonins. Molecular Cell 24(1):25-37

Reissmann, S., C. Parnot, C.R. Booth, W. Chiu, and J. Frydman (2007). Essential function of the built-in lid in the allosteric regulation of eukaryotic and archaeal chaperonins. Nat Struct Mol Biol 14: 432-40.

Chen, D. H., Song, J. L., Chuang, D. T., Chiu, W. & Ludtke, S. J. (2006). An expanded conformation of single-ring GroEL-GroES complex encapsulates an 86 kDa substrate. Structure 14: 1711-22.

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