1. Coiled Coils 7.88J Protein Folding Prof. David Gossard Room 3-336, x3-4465 Gossard@mit.edu September 28, 2005

2. Outline Key Features of Coiled Coils A Particular Example –GCN4 Leucine Zipper (2ZTA)

3. Fibrous protein examples Tropomyosin Intermediate filament protein Lamin M-protein Paramyosin Myosin Cohen, C. and D.A.D. Perry, (1990) “  -helical coiled coils and bundles: How to design an  -helical protein”, PROTEINS: Structure, Function, and Genetics 7:1-15.

4. Other example – GCN4 Gene regulation in yeast Recognizes a specific DNA sequence –  -helices’ sidechains contact major groove of DNA –DNA-protein “fit” is specific and strong Protein dimerization and DNA binding functions are integrated Alberts, et.al., (2002), Molecular Biology of the Cell, 4 th edition, Garland

5. Coiled Coils Left-handed spiral of right-handed helices May be parallel or anti-parallel N C N N N C C C

6. Equations of Helix (Coil) = tan –1 (2  r 0 /p 0 ) r o - radius P o - pitch  = pitch angle

7. Equations of Coiled-Coil x(t) = r 0 cos  0 t + r 1 cos  0 t cos  1 t - r 1 cos  sin  0 t sin  1 t y(t) = r 0 sin  0 t + r 1 sin  0 t cos  1 t + r 1 cos  cos  0 t sin  1 t z(t) = p 0 (  0 t) - r 1 sin  sin  1 t F.H.C. Crick, “The Fourier Transform of a Coiled-coil”, Acta Cryst. (1953), 6, 685-689  = tan –1 (2  r 0 /p 0 ) x y z

8. Questions What is the nature of the interaction between the coils? What is the angle of twist? What are the sequence determinants?

9. “Knobs in Holes” Packing F.H.C. Crick, “The Packing of  -helices: Simple Coiled-coils”, Acta Cryst. (1953), 6, 689-697 Helix axis

10. Features of Coiled Coil Heptad repeat in sequence –[a b c d e f g] n Hydrophobic residues at “a” and “d” Charged residues at “e” and “g” Hydrophobic residues at “a” and “d” Charged residues at “e” and “g” +/-

11. Significance of Heptad Repeat Figure adapted from Cohen, et.al., PROTEINS: Structure, Function and Genetics 7:1-15 (1990) Residues at “d” and “a” form hydrophobic core Residues at “e” and “g” form ion pairs +/- -/+

12. Heptad Repeat in 3D a b c d e f g a b c d e f g Hydrophobic residues Charged residues +/- -/+ +/- -/+

13. Hydrophobic Core is on Axis of Superhelix ( ~Straight) d ad a

14. Charged Residues Provide Stability, Registration Charged residues “e” and “g” Ion pairs between coils

15. Demonstration Heptad repeat in 3D Full Coiled Coil in 3D “Knobs in Holes” Packing

16. GCN4-p1 Leucine Zipper (2ZTA) Parallel Coiled Coil (last) 31 residues ~ 45 A ~ 8 turns Separation of minor axes ~ 9.3 A Major helix pitch ~ 181 A/turn Major helix ~ 90 o Erin O’Shea, Juli D. Klemm, Peter S. Kim, and Tom Alber, “X-ray Structure of the GCN4 Leucine Zipper, a Two-Stranded, Parallel Coiled Coil”, Science, 254, pp. 539-544, October 25, 1991

17. GCN4-p1 Leucine Zipper (2ZTA) Residues contain heptad repeat Ion pairs –Lys 15 – Glu 20’ –Glu 22 – Lys 27’ –Glu 22’ – Lys 27

18. Crossing Angle ~18 o

19. 3-Stranded Coiled Coil!? (parallel) Axial symmetry Hydrophobic core Ion pairs

20. 4-Stranded Coiled Coil!? (parallel) Axial symmetry Hydrophobic core Ion pairs

21. 3 & 4-Stranded Coiled Coils 3-stranded –Gp17 (T7) –Fibrinogen (heterotrimer) –GCN4 mutant 4-stranded parallel –GCN4 mutants 4-stranded anti-parallel –Myohaemerythrin –Tobacco mosaic virus –Cytochrome c’ –Apoferritin

22. END