4-chloroprolines: synthesis, conformational analysis, and effect on the collagen triple helix - PubMed (original) (raw)

4-chloroprolines: synthesis, conformational analysis, and effect on the collagen triple helix

Matthew D Shoulders et al. Biopolymers. 2008 May.

Abstract

Collagen is an abundant, triple-helical protein comprising three strands of the repeating sequence: Xaa-Yaa-Gly. (2S)-Proline and (2S,4R)-4-hydroxyproline (Hyp) are common in the primary structure of collagen. Here, we use nonnatural proline derivatives to reveal determinants of collagen stability. Specifically, we report high-yielding syntheses of (2S,4S)-4-chloroproline (clp) and (2S,4R)-4-chloroproline (Clp). We find that the molecular structure of Ac-Clp-OMe in the solid state is virtually identical to that of Ac-Hyp-OMe. In contrast, the conformational properties of Ac-clp-OMe are similar to those of Ac-Pro-OMe. Ac-Clp-OMe has a stronger preference for a trans amide bond than does Ac-Pro-OMe, whereas Ac-clp-OMe has a weaker preference. (Pro-Clp-Gly)(10) forms triple helices that are significantly more stable than those of (Pro-Pro-Gly)(10). Triple helices of (clp-Pro-Gly)(10) have stability similar to those of (Pro-Pro-Gly)(10). Unlike (Pro-Clp-Gly)(10) and (clp-Pro-Gly)(10), (clp-Clp-Gly)(10) does not form a stable triple helix, presumably due to a deleterious steric interaction between proximal chlorines on different strands. These data, which are consistent with previous work on 4-fluoroprolines and 4-methylprolines, support the importance of stereoelectronic and steric effects in the stability of the collagen triple helix and provide another means to modulate that stability. (

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Figures

FIGURE 1

FIGURE 1

Ring conformations of 4-substituted prolines. The Cγ -endo conformation is favored strongly by stereoelectronic effects when R1 = H, R2 = F (flp) or Cl (clp) and by steric effects when R1 = Me (mep), R2 = H. The Cγ -exo conformation is favored strongly by stereoelectronic effects when R1 = OH (Hyp), F (Flp) or Cl (Clp), R2 = H and by steric effects when R1 = H, R2 = Me (Mep).

FIGURE 2

FIGURE 2

(A) Molecular drawing of crystalline Ac–Clp–OMe (10; 50% probability ellipsoids). (B) Conformation of crystalline 10 and Ac-Hyp-OMe depicted with the program PyMOL (Delano Scientific, Palo Alto, CA).

FIGURE 3

FIGURE 3

Conformational analysis of clp- and Clp-containing CRPs. (A, C, and E) CD spectra of peptide solutions (0.2 mM in 50 mM HOAc) at 4 °C after incubating at ≤4 °C for ≥24 h. (B, D, and F) Effect of temperature on the molar ellipticity at 225 or 226 nm. Data were recorded at 3-°C intervals after a 5-min equilibration.

FIGURE 4

FIGURE 4

Space-filling models of segments of triple helices constructed from the three-dimensional structure of a (Pro-Hyp-Gly)n triple helix (PDB entry 1CAG11) by replacing the H or OH on Pro and Hyp with F or Cl, respectively, using the program SYBYL (Tripos, St. Louis, MO) and depicting the images with the program PyMOL. (A) Segment of a (flp-Flp-Gly)n triple helix (_r_F···F = 2.4 Å). (B) Segment of a (clp-Clp-Gly)n triple helix (_r_Cl···Cl = 1.9 Å).

SCHEME 1

SCHEME 1

SCHEME 2

SCHEME 2

SCHEME 3

SCHEME 3

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