No abstract

This new scoring system is a valid technique to assist in the diagnosis of constipation and is the first study using appropriate statistical methodology to demonstrate a discriminatory ability of multiple symptoms in constipation. At present, symptom analysis does not adequately differentiate major pathophysiologic subgroups for use in clinical practice.

Length-dependent helical propensities w Ala (n,T) at T = 10, 25, and 60 °C are assigned from t/c values and NMR 13 C chemical shifts for series 1 peptides TrpLys m Inp 2 t Leu-Ala n t LeuInp 2 Lys m NH 2 , n = 15, 19, and 25, m = 5, in water. Van't Hoff analysis of w Ala (n,T) show that α-helix formation is primarily enthalpy-driven. For series 2 peptides Ac-Trp Lys 5 Inp 2 t Leu-β AspHel-Ala nbeta-t LeuInp 2 Lys 5 NH 2 , n = 12 and 22, which contain exceptionally helical Ala n cores, protection factor-derived fractional helicities FH are assigned in the range 10-30 °C in water and used to calibrate temperature-dependent CD ellipticities [θ] λ,H,n,T . These are applied to CD data for series 1 peptides, 12 ≤ n ≤ 45, to confirm the w Ala (n,T) assignments at T = 25 and 60 °C. The [θ] λ,H,n,T are temperature dependent within the wavelength region, 222 ± 12 nm, and yield a temperature correction for calculation of FH from experimental values of [θ] 222,n,T,Exp .

Measured at 2 °C in water, NMR chemical shifts of 13 C═O labeled central alanine residues of peptides W-Lys 5 -t L 3 -Ala n -t L 3 -Lys 5 NH 2 , n = 9, 11, 13, 15, 19 and W-Lys 5 -t L 3 -a-Ala n -A-Inp-t L 2 -Lys 5 NH 2 (a = D-Ala; t L = tert-leucine; Inp = 4-carboxypiperidine) are used to assign j L t and c L t , the N-and C-terminal t L capping parameters and length-dependent values for w Ala (n), the alanine helical propensity for Ala n peptides. These parameters allow Lifson-Roig characterization of the stabilities of Ala n helices in water. To facilitate chemical shift characterization, different 13 C/ 12 C ratios are incorporated into specific Ala sites to code up to six residue sites per peptide. Large left/right chemical shift anisotropies are intrinsic to helical polyalanines, and a correcting L-R-based model is introduced. Capping parameters j L t = c L t lie in the range of 0.3 to 0.5; the t L residues are thus moderately helix-destabilizing. For helical conformations of lengths shorter than eight residues, assigned values for w Ala approach 1.0 but increase monotonically with length to a value of 1.59 for w Ala (19).

NMR and CD studies are reported for two length series of solubilized, spaced, highly helical polyalanines that are N-capped by the optimal helix stabilizer β Asp-Hel and C-capped by β-aminoalanine beta and that are studied in water at 2 °C, pH 1-8. NMR analysis yields a structural characterization of the peptide Ac β AspHelAla 8 betaNH 2 and selected members of one β AspHelAla n beta series. At pH > 4.5 the β AspHel cap provides a preorganized triad of carboxylate anion and two amide residues that is complementary to the helical polyalanine Nterminus. The C-terminal β-aminoalanine assumes a helix-stabilizing conformation consistent with literature precedents. H(N)CO NMR experiments applied to capped, uniformly 13 C-and 15 N-labeled Ala 8 and Ala 12 peptides define Ala n hydrogen bonding signatures as α-helical without detectable 3 10 character. Relative NH→ND exchange rates yield site protection factors PF i that define uniquely high fractional helicities FH for the peptide Ala n regions. These Ala n calibration series, studied in water and lacking helix-stabilizing tertiary structure, yield the first 13 C NMR chemical shifts, 3 J HNHα coupling constants, and CD ellipticities [θ Molar ] λ,n characteristic of a fully helical alanine within an Ala n context. CD data are used to assign parameters X and [θ] λ,∞ , required for rigorous calculation of FH values from CD ellipticities.