Lipid monolayer 2-D crystallization is a technique that was pioneered by Uzgiris and Kornberg (Uzgiris and Kornberg, 1983). It has been used for obtaining 3-D images, mostly in negative stain, of a number of proteins (Darst et al., 1991; Darst et al., 1989; Jontes and Milligan, 1997; Ribi et al., 1988; Stoilova-McPhie et al., 2002). Our work on lipid monolayer crystallization and structure analysis in unique in that it utilizes only unstained frozen hydrated 2-D arrays for 3-D imaging. Some of these examples are shown below.
For this structure work, we use a method called hybrid crystallography. This involves obtaining a 3-D image at less than atomic resolution and interpreting it using higher resolution structures docked into the 3-D maps as rigid bodies.
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Taylor, K.A., D.W. Taylor, and F. Schachat, Isoforms of alpha-actinin from cardiac, smooth, and skeletal muscle form polar arrays of actin filaments. J. Cell Biol., (2000). 149(3), 635-46. (Pubmed Link)
Our laboratory has been working on the structure of alpha-actinin since 1993 when we produced the first 2-D arrays of this protein. Initial efforts concentrated on chicken smooth muscle alpha-actinin, then expanded to rabbit skeletal muscle isoforms with current efforts concentrating on non-muscle isoforms. alpha-actinin crystallizes readily in 2-D arrays on a lipid monolayer making electron c an ideal method for structure analysis.
Tang, J., D.W. Taylor, and K.A. Taylor, The three-dimensional structure of alpha-actinin obtained by cryoelectron microscopy suggests a model for Ca(2+)-dependent actin binding. J Mol Biol, (2001). 310(4), 845-58. (Pubmed Link)
Liu, J., D.W. Taylor, and K.A. Taylor, A 3-D reconstruction of smooth muscle alpha-actinin by cryoEM reveals two different conformations at the actin binding region. J. Mol. Bio., (2004). 338(1), 115-125. (Pubmed Link)
Wendt, T., D. Taylor, T. Messier, K.M. Trybus, and K.A. Taylor, Visualization of head-head interactions in the inhibited state of smooth muscle myosin. J. Cell. Biol., (1999). 147(7), 1385-90. (Pubmed Link)
Wendt, T., D. Taylor, K.M. Trybus, and K. Taylor, Three-dimensional image reconstruction of dephosphorylated smooth muscle heavy meromyosin reveals asymmetry in the interaction between myosin heads and placement of subfragment 2. Proc. Natl. Acad. Sci. U S A, (2001). 98(8), 4361-6. (Pubmed Link)
Liu, J., T. Wendt, D. Taylor, and K. Taylor, Refined model of the 10S conformation of smooth muscle myosin by cryo-electron microscopy 3D image reconstruction. J. Mol. Biol., (2003). 329(5), 963-72. (Pubmed Link)