Applications in Structural Biology

To determine the potential of nanotube tips in structural biology, DNA and several well-characterized proteins have been imaged. DNA was imaged by manually assembled MWNT tips in air [79] and in fluid [85]. The fluid imaging experiments produced a measured height of 2 nm, the expected value based on the intrinsic DNA diameter, and the resolution in these studies was on the order of 3.5 to 5nm. These values for the resolution are consistent with that expected for multiwalled nanotube material, but are also similar to the best values observed with microfabricated tips [86,72].

Studies of isolated proteins provide a more stringent test of the capabilities of probe tips, and demonstrate clearly the advantages of nanotube tips. Pore-growth MWNT CVD tips have been used to image isolated immunoglobulin-M (IgM) antibody proteins [68]. IgM is a ca. 1 MDa antibody protein with a pentameric structure. It has not been crystallized for X-ray diffraction, but electron microscopy has elucidated the basic features of the pentameric structure [87]. Room temperature studies with pore-growth CVD MWNT nanotube tips [68] have clearly shown the pentameric structure, including five external pairs of antigen binding fragments (Fab domains) and five internal Fc fragments (Fig. 21).

Fig. 21. Images of immunoglobulin-M (IgM) taken by CVD MWNT tips. Many IgM molecules are seen in various conformations due to their inherent flexibility. Scale bar is 100 nm. The inset shows an example of a well-oriented IgM molecule with the potential J-loop structure highlighted by the white arrow. Scale bar is 10 nm [68]

In addition, these images occasionally exhibited a structure connecting two of the five Fc domains that could correspond to the joining (J) loop [68]. Because the exact structure of this region is still unclear [87], these investigations have shown the potential of nanotube probes to reveal new structural features on large proteins that cannot be crystallized for diffraction studies.

The resolution of the smaller diameter CVD SWNT tips has been further tested in studies of a smaller protein, GroES, which is a component of the GroEL/GroES chaperoning system. GroES is a hollow dome shaped hep-tamer that is approximately 8nm in outer diameter [88]. The seven 10 kD subunits each consist of a core b-barrel with a b-hairpin loop at the top and bottom. The top b-hairpins point inward to form the top of the dome, while the bottom hairpins are disordered when not in contact with GroEL [89]. CVD SWNT tip images of individual, well-separated GroES molecules on mica [82] reveal that it is possible to resolve the seven-fold symmetry [89] as shown in Fig. 22. These results have demonstrated clearly the ability of the present CVD nanotube tips to achieve sub-molecular resolution on isolated protein systems.

Fig. 22. GroES images taken by CVD SWNT tips. (a) A field of many GroES are shown, some displaying the pore side and some displaying the dome. Scale bar is 50 nm. (b) A high-resolution image showing the heptameric structure. (c) The crystal structure for (b) is shown for comparison. [82] Scale bar in (b) and (c) is 5 nm

Fig. 22. GroES images taken by CVD SWNT tips. (a) A field of many GroES are shown, some displaying the pore side and some displaying the dome. Scale bar is 50 nm. (b) A high-resolution image showing the heptameric structure. (c) The crystal structure for (b) is shown for comparison. [82] Scale bar in (b) and (c) is 5 nm

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