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Modeling of protein turns and derivation of NMR parameters related to turn structure Megan Chawner BRITE REU Program Advisor: Dr. Dimitrios Morikis Department.

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Presentation on theme: "Modeling of protein turns and derivation of NMR parameters related to turn structure Megan Chawner BRITE REU Program Advisor: Dr. Dimitrios Morikis Department."— Presentation transcript:

1 Modeling of protein turns and derivation of NMR parameters related to turn structure Megan Chawner BRITE REU Program Advisor: Dr. Dimitrios Morikis Department of Bioengineering University of California, Riverside

2 Outline Background My Project Results Conclusions Acknowledgements

3 Protein Structure: All proteins are made up of twenty amino acid building blocks into a sequence = primary structure

4 Protein structure: sequence folds into  -sheet,  -helix, random coil loops and various types of turns stabilized by atomic interactions (e.g., H-bonds) = secondary structure Anti-parallel  -sheet  -helix Primary structure: GPLLNKFLTT Primary structure: EKQKPDGVFQE Strand 1 Strand 2 Inter-strand H-bonds C=O(i)…H-N(i+4) H-bonds 1 helix turn = 3.6 a.a.

5 Protein Structure: three-dimensional protein folds are stabilized by long range interactions = tertiary structure Turns introduce reversibility in the direction of other elements of secondary structure, such as  -helices or  -sheets 3 amino acids =  -turn 4 amino acids =  -turn  -turn  -turn

6 i-1 i i+1 ii ii ii  -sheet Ramachandran plot (  ) plot defines secondary structure Backbone torsion angles:  Turns  -helix

7 Protein Structure Determination: uses Nuclear magnetic resonance (NMR) spectroscopy to get NMR observables, which are converted to NMR-derived structural parameters Nuclear Overhauser effects (NOEs)  inter-proton distances 3 J(H N -H  )-coupling constants   -torsion angles Karplus Equation (Karplus, 1959, J Chem Phys) NOE equation (Wuthrich, 1986) r i,j  inter-proton distance  c  rotational correlation time A=6.98, B=-1.38, C=1.72 (Wang and Bax, 1996, JACS) NOE < 5 Å  through-space interactions  inter-proton distances 3 J(H N -H  ) = 3-chemical bond coupling  through-bond interactions   -torsions

8 Amino Acid iAmino Acid i+1 NCC C OR H NCC C O H R ii ii 3 J(H N -H  )  i+1  i+1 HH ii H N (i)-H  (i) H N (i)-H N (i+1) H N (i)-H  (i+1) 3 J(H N -H  ) = 3-bond   -torsion NOE < 5 Å  distance in space H  (i)-H  (i+1) H  (i)-H N (i+1) Relations of experimental observables and structural parameters d  N (i,i+1) d NN (i,i+1) d N  (i,i) d  (i,i+1)

9 3 J(H N -H  ) (Hz)  ( o ) Cis  =0 o  =60 o  =90 o  =150 o Newman Projections N C=O C=O H H C  N C=O C=O H H C  N C=O C=O H H C  N C=O C=O H H C   =-90 o  =-30 o Trans  =180 o  =-120 o Solution of Karplus equation using MatLab  -helix  -sheet NC  H  H Cis NC  H  H Trans NC HH Cis NC  H  H Trans C NC  C  Cis C NC  C  Trans C NC  C  Cis C NC  C  C NC  C  C NC  C  Trans C NC  C  C NC  C  Chawner & Morikis, in preparation

10 My Project Goals: To use NMR-derived parameters (inter-proton distances and  -torsion angles) to create databases of expected NMR observables (NOEs and 3 J(H N -H  )- coupling constants) for ideal  - and  - turns with statistical deviations. Bottom line: we are back-calculating NMR observables. Remember, during structure determination, NMR-derived parameters are obtained from NMR spectroscopic observables, NOEs and 3 J(H N -H  )-coupling constants. Use: Rapid protein turn structure identification by examination of raw NMR observables, without a complete structure calculation.

11 Color code: Blue: N Light blue: H Gray: C Red: O Color code: Blue: N Light blue: H Gray: C Red: O VIII I I’ II’ II 1 3 2 4         H-bond C  -C  I I’ II’ II 1 3 2 4         H-bond C  -C   -turns Computational modeling of ideal  -and  -turns according to torsion angles using DeepView Classic  -turn criteria Distance: C  (1)-C  (4) < 7 Å C=O(1)…H-N(4) H-bonded Distance: O(1)-N(4) < 3.3 Å Distance: O(1)-HN(4) < 2.4 Å Angle: O(1)-H(4)-N(4) almost linear ± 35 o Torsion angles ( o ) Type 22 22 33 33 I-60-30-900 II-60120800 I'6030900 II'60-120-800 VIII-60-30-120120 Chawner & Morikis, in preparation

12 Torsion angles ( o ) Type 22 22 Direct70-60 70-70 85-60 85-70 Inverse-7060 -7070 -8560 -8570 directinverse  -turns Computational modeling of ideal  -and  -turns according to torsion angles Classic  -turn criteria Chawner & Morikis, in preparation

13 Nuclear Overhauser effects (NOEs)  inter-proton distances Characteristic  -turn distances H  (2)-H N (4): (i, i+2) H  (2)-H N (3): (i, i+1) H  (3)-H N (4): (i, i+1) H N (2)-H N (3): (i, i+1) H N (3)-H N (4): (i, i+1) Characteristic  -turn distances H  (1)-H N (3): (i, i+2) H  (1)-H N (2): (i, i+1) H  (2)-H N (3): (i, i+1) H N (1)-H N (2): (i, i+1) H N (2)-H N (3): (i, i+1)  -turn  -turn

14 Torsion angles ( o )D < 7 ÅH-bond distance (Å)H-bond angle (°) Type 22 22 33 33 C  (1)-C  (4)O(1)-N(4)O(1)-H N (4)O(1)-H(4)-N(4) I-60-30-9004.72.61.6153.2 II-601208004.72.61.7152.2 I'60309004.73.02.1151.1 II'60-120-8004.72.92.0153.4 VIII-60-30-1201206.24.34.569.5 Torsion angles ( o )D < 7 ÅH-bond distance (Å)H-bond angle (°) Type 22 22 C  (1)-C  (3)O(1)-N(3)O(1)-H N (3)O(1)-H(3)-N(3) Direct70-605.42.71.8142.2 70-705.52.71.9135.5 85-605.53.12.2137.4 85-705.63.12.3134.6 Inverse-70605.42.41.5143.6 -70705.52.51.7131.3 -85605.52.81.9141.9 -85705.62.82.0136.0 Marginal H-bonds present because of larger deviations from linearity Test of compliance of molecular models with ideal turn criteria Not present H-bond present Chawner & Morikis, in preparation

15 Inter-proton distance (Å) Type H N (2)- H N (3) H N (3)- H N (4) H  (2)- H N (3) H  (3)- H N (4) H  (2)- H N (4) H  (2)- H  (3) H  (3)- H  (4) H N (2)- H  (3) H N (3)- H  (4) I2.62.43.53.33.74.74.85.34.7 II4.52.52.13.3 4.44.86.45.2 I'2.62.43.03.34.24.8 5.0 II'4.52.53.3 4.24.54.85.74.9 VIII2.64.33.52.15.84.64.45.34.9 Torsion angles (°) Inter-proton distance (Å) Type 22 22 H N (1)- H N (2) H N (2)- H N (3) H  (1)- H N (2) H  (2)- H N (3) H  (1)- H N (3) H  (1)- H  (2) H  (2)- H  (3) H N (1)- H  (2) H N (2)- H  (3) Direct70-602.03.73.6 4.05.34.83.95.7 70-702.03.83.6 4.25.34.73.95.7 85-602.03.6 4.25.34.83.85.5 85-702.03.83.6 4.45.34.73.85.6 Inverse-70602.03.73.62.63.84.84.64.55.1 -70702.03.83.62.54.14.84.64.55.1 -85602.03.6 2.63.94.74.64.44.9 -85702.03.83.62.54.24.74.64.44.9 Ideal  -turns Ideal  -turns Molecular models: measured distances corresponding to characteristic NOEs

16 We classified the inter-proton distances as corresponding to strong, medium, weak and very weak NOE intensities: 1.8-2.6 Å = strong 2.7-3.5 Å = medium 3.6-4.4 Å = weak 4.5-5.0 Å = very weak Relative NOE intensities Type H N (2)- H N (3) H N (3)- H N (4) H  (2)- H N (3) H  (3)- H N (4) H  (2)- H N (4) H  (2)- H  (3) H  (3)- H  (4) H N (2)- H  (3) H N (3)- H  (4) ISSMMWVW N/OVW IIVWSSMMW N/O I'SSMMWVW II'VWSMMW N/OVW VIIISWMSN/OVWWN/OVW  -turns Relative classification of NOE intensities Chawner & Morikis, in preparation 1.8 Å: sum of van der Waals radii with some overlap

17 Torsion angles (°) Relative NOE intensities Type 22 22 H N (1)- H N (2) H N (2) - H N (3) H  (1) - H N (2) H  (2) - H N (3) H  (1) - H N (3) H  (1) - H  (2) H  (2) - H  (3) H N (1) - H  (2) H N (2) - H  (3) Direct70-60SWWWWN/OVWWN/O 70-70SWWWWN/OVWWN/O 85-60SWWWWN/OVWWN/O 85-70SWWWWN/OVWWN/O Inverse-7060SWWSWVW N/O -7070SWWSWVW N/O -8560SWWSWVW W -8570SWWSWVW W  -turns We classified the inter-proton distances: 1.8-2.6 Å = strong 2.7-3.5 Å = medium 3.6-4.4 Å = weak 4.5-5.0 Å = very weak Relative classification of NOE intensities

18  2 (°) J 2 (Hz)  3 (°) J 3 (Hz) Type I-604.2-908.2 Type I’607.3905.8 Type II-604.2806.6 Type II’607.3-806.9 Type VIII-604.2-12010.1 Type  2 (°) J 2 (Hz) Direct707.1 Direct856.2 Inverse-705.5 Inverse-857.5 Solution of Karplus equation: calculations of characteristic 3 J(H N -H  )-coupling constants  -turns  -turns Chawner & Morikis, in preparation

19  2 (°) J 2 (Hz)  3 (°) J 3 (Hz) Type I-60Weaker-90Stronger Type I’60Stronger90Weaker Type II-60Weaker80Stronger Type II’60Stronger-80Weaker Type VIII-60Weaker-120Stronger Type  2 (°) J 2 (Hz) Direct70S Direct85W Inverse-70W Inverse-85S We classified the turn’s 3 J(H N -H  )-coupling constants as stronger or weaker relative to itself, so that the different types can be differentiated comparatively  -turns  -turns Caution: small variations in  -torsion angles result to very large variations in j-coupling constants. In general, the use of j-coupling constants is not as helpful as NOE intensity patterns and connectivities.  -helix  -sheet Chawner & Morikis, in preparation

20 Conclusions NOE intensity patterns and connectivities can be used to distinguish turn type without a complete structure determination. We have created small NOE intensity databases that discriminate Type I, I’, II, II’, and VIII  -turns, and direct and inverse  -turns.  Caution: Classification of strong, medium, weak, and very weak NOEs is relative. Small variations of the characteristic  -torsion angles introduce very large variations in the 3 J(H N -H  )-coupling constant values, sometimes spanning the whole range of possible solutions for the Karplus equation and the whole allowed region of the Ramachandran plot.  Why? the small variations in  -torsion angles are owed to the dynamic character of turns in proteins and peptides and to conformational averaging. Overall, NOEs are more useful than J-coupling constants.

21 Acknowledgements Dr. Dimitrios Morikis Li Zhang Coordinators of BRITE Program Fellow BRITE students

22 3 J(H N -H  )  Cis  =0 o  =60 o  =90 o  =150 o Newman Projection N C=O C=O H H C  N C=O C=O H H C  N C=O C=O H H C  N C=O C=O H H C   =-90 o  =-30 o Trans  =180 o  =-120 o Solution of Karplus equation

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