Our objective is to determine the precise molecular architecture of certain alpha-fibrous proteins that have dynamic as well as structural roles in the cell. The chief methods are coordinated electron microscopy and X-ray crystallography, together with biochemistry. Muscle proteins are a central focus and provide the background for studying related systems. A major aim is t determine the structure, motions and interactions in three protein switches that control contraction. The topology of the myosin head and its associated light chains, the crystal structure of the tropomyosin/troponin complex, and the structure and interactions of the myosin rod and paramyosin are being analyzed to establish models for regulation. In each case, the structure of the individual protein components will be determined to the highest possible resolution, and the changes in structure and interactions that accompany switching will be sought. The molecular basis of blood clotting is being studied by a similar approach: the crystal structure of fibrinogen is being determined by X-ray crystallography and cryoelectron microscopy in order to understand its self-assembly into the fibrin clot. The principle of self-assembly applies to many of these proteins. This means that under appropriate conditions, these highly organized structures can be dissociated and reassembled in vitro to form ordered structures (including crystals) closely related to those found in vivo. In contrast to the ordered arrays formed by the purified proteins, however, the native structures are not so accessible to detailed analysis. The alpha-helical coiled-coil motif that characterizes tropomyosin, myosin rod and paramyosin has been shown to have a widespread occurrence in a diverse range of proteins. Using predictive methods based on known structures, we aim to define more fully the designs of other related alpha- proteins (such as dystrophin, spectrin,land alpha-actinin) which have not yet been solved crystallographically. Knowledge of the molecular mechanism of contraction and its regulation is needed to account for malfunctions in various heart and muscle pathologies. Similarly, a full understanding of blood clotting, and its malfunction in certain cardiovascular diseases, requires detailed information about the structure and interactions of the fibrinogen molecule. These studies aim to identify distinctive features of fibrous protein structure that are essential to function, and will lead to a deeper understanding of both normal and abnormal cell functions.

Agency
National Institute of Health (NIH)
Institute
National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIAMS)
Type
Method to Extend Research in Time (MERIT) Award (R37)
Project #
5R37AR017346-21
Application #
2078363
Study Section
Biophysical Chemistry Study Section (BBCB)
Project Start
1974-09-01
Project End
1995-02-28
Budget Start
1994-03-01
Budget End
1995-02-28
Support Year
21
Fiscal Year
1994
Total Cost
Indirect Cost
Name
Brandeis University
Department
Biology
Type
Schools of Arts and Sciences
DUNS #
616845814
City
Waltham
State
MA
Country
United States
Zip Code
02454
Gillilan, Richard E; Kumar, V S Senthil; O'Neall-Hennessey, Elizabeth et al. (2013) X-ray solution scattering of squid heavy meromyosin: strengthening the evidence for an ancient compact off state. PLoS One 8:e81994
O'Neall-Hennessey, Elizabeth; Reshetnikova, Ludmila; Senthil Kumar, V S et al. (2013) Purification, crystallization and preliminary X-ray crystallographic analysis of squid heavy meromyosin. Acta Crystallogr Sect F Struct Biol Cryst Commun 69:248-52
Brown, Jerry H (2013) Deriving how far structural information is transmitted through parallel homodimeric coiled-coils: a correlation analysis of helical staggers. Proteins 81:635-43
Kumar, V S Senthil; O'Neall-Hennessey, Elizabeth; Reshetnikova, Ludmila et al. (2011) Crystal structure of a phosphorylated light chain domain of scallop smooth-muscle myosin. Biophys J 101:2185-9
Brown, Jerry H; Kumar, V S Senthil; O'Neall-Hennessey, Elizabeth et al. (2011) Visualizing key hinges and a potential major source of compliance in the lever arm of myosin. Proc Natl Acad Sci U S A 108:114-9
Rosenbluth, Jack; Szent-Gyorgyi, Andrew G; Thompson, Joseph T (2010) The ultrastructure and contractile properties of a fast-acting, obliquely striated, myosin-regulated muscle: the funnel retractor of squids. J Exp Biol 213:2430-43
Brown, Jerry H (2010) How sequence directs bending in tropomyosin and other two-stranded alpha-helical coiled coils. Protein Sci 19:1366-75
Himmel, Daniel M; Mui, Suet; O'Neall-Hennessey, Elizabeth et al. (2009) The on-off switch in regulated myosins: different triggers but related mechanisms. J Mol Biol 394:496-505
Yang, Yuting; Gourinath, S; Kovacs, Mihaly et al. (2007) Rigor-like structures from muscle myosins reveal key mechanical elements in the transduction pathways of this allosteric motor. Structure 15:553-64
Szent-Gyorgyi, Andrew G (2007) Regulation by myosin: how calcium regulates some myosins, past and present. Adv Exp Med Biol 592:253-64

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