Theoretical π-π* absorption and circular dichroic spectra of β-turn model peptides

The dipole interaction model, treated by the partially dispersive normal mode method, is used to calculate π-π* absorption and circular dichroic spectra of β-turn model peptides in certain conformations. These include Ac-Gly-Gly-NHMe, Ac-L-Ala-L-Ala-NHMe, and Ac-L-Ala-Gly-NHMe in the standard β-turn conformations I, II, and III of Venkatachalam and cyclo(L-Ala-Gly-ε-aminocaproyl), cyclo(L-Ala-L-Ala—aminocaproyl), and cyclo(L-Ala-D-Ala-ε-aminocaproyl) in the minimum-energy conformations of Nemethy et al. Boltzmann average circular dichroic spectra of the cyclic compounds agree with experimental spectra in most respects. The results are compared with previous theoretical CD spectra for these molecules and with conformational assignments based on other evidence. Absorption spectra in the π-π* band are predicted to be moderately sensitive to conformation.     

Conformational investigation of α,β-dehydropeptides

Conformations of three series of model peptides: homochiral Ac-Pro-L-Xaa-NHCH₃ and heterochiral Ac-Pro-D-Xaa-NHCH₃ (Xaa=Phe, Val, Leu. Abu. Ala) as ivell as α,β-dehydro Ac-Pro-ΔXaa-NHCH_s [ΔXaa = (Z)-ΔPhe, ΔVal. (Z)-ΔLeu, (Z)-ΔAbu] were investigated by CD spectroscopy in 2 % dichloromethanecyclohexane, trifluoroethanol. water. and occasionally in other solvents. The spectra of homochiral peptides show a significant solvent dependence. Folded structures are present in 2% dichloromethane-cyclohexane and unordered ones occur in water. The folded conformers are of the inverse γ-turn type for all the peptides but Ac-Pro-L-Phe-NHCH₃ for which the type-I β-turn is preferred. The changes in the spectra of the heterochiral peptides are limited. The compounds adopt the typc-II β–turn in 2% dichloromethanecyclohexane, represented by class B spectra, and retain this conformation in water as well as in fluorinated alcohols but not always to a full extent. The CD spectra of the unsaturated peptides in 2%, dichloromethanecyclohexane, although they cannot be assigned to any common spectral class, must be attributed to the βII-turn conformation as determined for these coinpounds by NMR and IR spectroscopy. The CD spectra of dehydropeptides exhibit a considerable solvent dependence and suggest unordered structures in water.     

Dipole interaction model predicted π−π* circular dichroism of cyclo(l‐Pro)3 using structures created by semi‐empirical,ab initio,and molecular mechanics methods

Abstract: Cyclo(l ‐Pro)₃ (CP3) is a synthetic peptide created to model cis and torsionally strained peptide bonds that also exhibits a strong distinctive UV circular dichroic (CD) spectrum. Circular dichroic spectra were computed for the amide π ? π* transition using the dipole interaction model for various conformations of the peptide. Conformations of CP3 were created initially from crystal data, and followed by energy minimizations via molecular mechanics using the cvff force field; the effects of additional geometric optimizations by semi‐empirical and ab initio quantum mechanics were investigated. The CD spectra for each conformation were calculated using a variety of different parameters, and each result was compared with the published experimental spectrum [Deber, C.M., Scatturin, A., Vaidya, V.M. & Blout, E.R. (1970) Small cyclic proline peptides: UV absorption and CD. In: Peptides: Chemistry and Biochemistry, Proceedings of the First American Peptide Symposium (Weinstein, B., ed.), Marcel Dekker, New York pp. 163–173]. Herein, two distinct conformations, a C₃ symmetric and an asymmetric form, gave CD predictions that separately did not resemble the experimental spectrum. Energy differences were predicted at various theoretical levels, including MP2 and density functional theory. When the predicted CD spectra for each conformation were multiplied by Boltzmann weighting factors created using heats of formation determined by the AM1 optimizations, the weighted composite CD spectrum created did resemble experiment for the π ? π* transition indicating that both conformations may exist simultaneously in solution.     

Conformations of ψ[CH2NH] pseudopeptides

The cyclic pseudopentapeptide cyclo[Gly-Proψ[CH₂NH]Gly-D-Phe-Pro] and its TFA salt were synthesized by solution methods, and their conformations were studied by NMR spectroscopy in both DMSO-d₆ and CDC₁₃. While intramolecular hydrogen bonding is observed with some conformers, the nature of the solvent and the presence of TFA affects the relative structural rigidities of the compounds. No evidence was found for the ψ[CH₂NH] or ψ[CH₂NH⁺₂] units acting as H donors in this series.     

The evaluation of type I and type II β-turn mixtures

Circular dichroism (CD) and¹ H-{¹H}NOE spectra were obtained for Piv-Pro-Ser-NHCH₃(1),[Piv-(CH₃)₃-C-CO], Boc-Pro-Ser-NHCH₃ (2) and Boc-Val-Ser-NHCH₃ (3), to determine the solution conformation of these p-turn models. In the crystal, 1 and 3 adopt an ideal type I β-turn, while 2 is characterized by a semifolded backbone geometry incorporating a cis Boc-Pro tert-amide bond. The predominance of a β-turn conformation in solution was suggested for models 1-3 on the basis of ¹H-{¹H}NOE data. In a nonpolar solvent the prevailing trans rotamer form (>80%) of 2 has a β-turn conformation according to heteronuclear NOE measurement. Positive ¹H-{¹H} NOEs were detected between the H^α(Pro)/NH(Ser), Hα(Ser)/NH(Ser) and NH(NHCH₃3)/HN(Ser) protons in the trans Boc-Pro rotamer form of 2 at -20° in CDCl₃. Similar positive homonuclear NOE enhancements were also observed on the appropriate proton signals in other models, such as Boc-Val-Ser-NHCH₃ (3). Boc-Val-D-Ser-NHCH₃ (4) and Boc-Pro-D-Ser-NHCH₃ (5), in various solvents. The ¹H- {¹H)NOE experiments carried out in CD₃CN clearly showed that besides the type I (or III) β-turn structure, one of the main conformations of models 1-5 is close to the type II β-turn backbone geometry in a nonpolar solvent. Unexpectedly, the conformational mixture of models 1-3 were characterized by class C (helix-like) CD spectra, although class C spectra are generally only correlated with the type I β-turn conformation. These acyclic models are the first carefully investigated examples of -L-L- triamide systems, containing a significant amount of a type II β-turn, as well as the type I p-turn and, however, yielding a class C circular dichroism spectra. The CD spectra recorded for 3 and 4 in acetonitrile were ‘calibrated’ using the ¹H-{¹H}NOE data. Such a “calibration”, as well as the semi-quantitative CD and NMR comprehensive analyses, demonstrated that class C, class B, as well as class C’ CD spectra may be obtained from the linear combination of the same two-component spectra, with different conformational weights. Therefore, it is suggested that the extraction of the conformational components of such models, simply on the basis of their CD spectra, must be made with caution.     

Conformational investigation of α,β-dehydropeptides VII*. Conformation of Ac-Pro-ΔAla-NHCH3 and Ac-Pro-(E)-ΔAbu-NHCH3: comparison with (Z)-substituted α,β-dehydropeptides†

The crystal structure and solution conformation of Ac-Pro-ΔAla-NHCH₃ and the solution conformation of Ac-Pro-(E)-ΔAbu-NHCH₃ were investigated by X-ray diffraction method and NMR, FTIR and CD spectroscopies. Ac-Pro-ΔAla-NHCH, adopts an extended-coil conformation in the crystalline state, with all-trans peptide bonds and the ΔAla residue being in a C₅ form, φ₁=– 71.4(4), ψ₁=– 16.8(4), φ₂=– 178.4(3) and ψ₂= 172.4(3)°. In inert solvents the peptide also assumes the C₅ conformation, but a γ-turn on the Pro residue cannot be ruled out. In these solvents Ac-Pro-(E)- ΔAbu-NHCH₃ accommodates a βII-turn, but a minor conformer with a nearly planar disposition of the CO—NH and C=C bonds (φ₂～0°) is also present. Previous spectroscopic studies of the (Z)-substituted dehydropeptides Ac-Pro-(Z)- ΔAbu-NHCH, and Ac-Pro-ΔVal-NHCH₃ reveal that both peptides prefer a βII-turn in solution. Comparison of conformations in the family of four Ac-Pro-ΔXaa-NHCH₃ peptides let us formulate the following order of their tendency to adopt a β-turn in solution: (Z)- ΔAbu > (E)- δAbu > ΔVal; ΔAla does not. None of the folded structures formed by the four compounds is stable in strongly solvating media. © Munksgaard 1996.     

Conformational investigation of αβ-dehydropeptides

Solution conformations of three series of model peptides, homochiral Ac-Pro-L-Xaa-NHCH₃ and heterochiral Ac-Pro-D-Xaa-NHcH₃ (Xaa = Val, Phe, Leu, Abu. Ah) as well as αβ-unsaturated Ac-Pro-ΔXaa-NHCH₃ [Δ Xaa =ΔVal, (Z)-ΔPhe, (Z)-ΔLeu, (Z)-ΔAbu] were investigated in CDCl₃ and CH₂Cl₂ by ¹H-, ¹³C-NMR, and FTIR spectroscopy. NH stretching absorption spectra, solvent shifts Δδ for NH (Xaa) and NHCH₃ on going from CDCl₃ to (CD₃)₂SO, diagnostic interresidue proton NOEs, and trans-cis isomer ratios were examined. These studies performed showed the essential difference in conformational propensities between homochiral peptides (L-Xaa) on the one hand and heterochiral (D-Xaa) and αβ-dehydropeptides (ΔXaa) on the other. Former compounds are conformationally flexible with an inverse γ-bend, a β-turn, and open forms in an equilibrium depending on the nature of the Xaa side chain. Conformational preferences of heterochiral and αβ-dehydropeptides are very similar, with the type-II β-turn as the dominating structure. There is no apparent correlation between conformational properties and the nature of the Xaa side chain within the two groups. The β-turn formation propensity seems to be somewhat greater in αβ-unsaturated than in heterochiral peptides, but an estimation of β-folded conformers is risky.     

Peptide models for β-turns.

The circular dichroism spectra of four β-turn model peptides, Z-Aib-Pro-Aib-Pro-OMe (1), Piv-Pro-Aib-NHMe (2), Piv-Pro-D-Ala-NHMe (3) and Piv-Pro-Val-NHMe (4) have been examined under a wide range of solvent conditions, using methanol, hexafluoroisopropanol and cyclohexane. Type I and Type II β-turns have been observed for peptides 1 and 2 respectively, in the solid state, while the Pro-D-Ala sequence adopts a Type II β-turn in a related peptide crystal structure. A class C spectrum is observed for 1 in various solvents, suggesting a variant of a Type I (III) structure. The Type II β-turn is characterized by a CD spectrum having two positive CD bands at ? 230 nm and ? 202 nm, a feature observed in Piv-Pro-D-Ala-NHMe in cyclohexane and methanol and for Piv-Pro-Aib-NHMe in methanol. Peptide 2 exhibits solvent dependent CD spectra, which may be rationalized by considering Type II, III and V reverse turn structures. Piv-Pro-Val-NHMe adopts non-β-turn structures in polar solvents, but exhibits a class B spectrum in cyclohexane suggesting a population of Type I β-turns.     

Design of peptides withα,β-dehydro-residues: synthesis,and crystal and molecular structure of a310-helical tetrapeptide Boc-l-Val-ΔPhe-ΔPhe-l-Ile-OCH3

Abstract: The peptide Boc-l -Val-ΔPhe-ΔPhe-l -Ile-OCH₃ was synthesized using the azlactone method in the solution phase, and its crystal and molecular structures were determined by X-ray diffraction. Single crystals were grown by slow evaporation from solution in methanol at 25°C. The crystals belong to an orthorhombic space group P2₁2₁2₁ with a = 12.882(7) Å, b = 15.430(5) Å, c = 18.330(5) Å and Z = 4. The structure was determined by direct methods and refined by a least-squares procedure to an R-value of 0.073. The peptide adopts a right-handed 3₁₀-helical conformation with backbone torsion angles: φ₁ = 56.0(6)°, ψ₁ = –38.0(6)°, φ₂ = –53.8(6)°, ψ₂ = 23.6(6)°, φ₃ = –82.9(6)°, ψ₃ = –10.6(7)°, φ₄ = 124.9(5)°. All the peptide bonds are trans. The conformation is stabilized by intramolecular 4→1 hydrogen bonds involving Boc carbonyl oxygen and NH of ΔPhe³ and CO of Val¹ and NH of Ile⁴. It is noteworthy that the two other chemically very similar peptides: Boc-Val-ΔPhe-ΔPhe-Ala-OCH₃ (i) and Boc-Val-ΔPhe-ΔPhe-Val-OCH₃ (ii) with differences only at the fourth position have been found to adopt folded conformations with two overlapping β-turns of types II and III′, respectively, whereas the present peptide adopts two overlapping β-turns of type III. Thus the introduction of Ile at fourth position in a sequence Val-ΔPhe-ΔPhe-X results in the formation of a 3₁₀-helix. The crystal structure is stabilized by intermolecular hydrogen bonds involving NH of Val¹ and carbonyl oxygen of a symmetry related (–x, y – 1/2, 1/2 + z) ΔPhe² and NH of ΔPhe² with carbonyl oxygen of a symmetry related (x, y1/2, 1/2 + z) Ile⁴. This gives rise to long columns of helical molecules linked head to tail running along [010] direction.     

Conformation of model,alanine and proline containing tetrapeptides in water

CD spectra of model alanine and prolyl-alanine tetrapeptides were measured at different _pH values. An analysis of the spectra shows that proline in position 2 or 4 of a tetrapeptide favours folding of the peptide chain, and unfolding when it is in position 3. Changes in CD spectra evidence growing amounts of the β-turn conformation upon increasing ^pH, independent of proline position in the peptide chain.     

Effect of the environment and role of the π-π stacking interactions in the stabilization of the 310-helix conformation in dehydroalanine oligopeptides

A quantum-mechanical study of the chain-length dependent stability of the extended, 2₇ribbon and 3₁₀-helix conformations in dehydroalanine (ΔAla) oligopeptides has been performed. To address the study, the oligopeptides ΔAla_n, where n varies from 1 to 6, were computed by using the semiempirical AM1 methodology. Cooperative free-energy effects permit one to predict the stabilization of the 3₁₀-helix with respect to the extended and 2₇-ribbon conformations when the number of residues in the polypeptide chain increases. The interactions associated with the π-electron density of the side chains can easily explain this finding. The effects of the solvent and the crystalline packing on the different conformations were modeled using a self-consistent reaction field (SCRF) method and a molecular mechanics approach to the packing, respectively. Both the aqueous and crystal environments seem to be a key factor in the stabilization of the helical conformation. Finally, the variations of electrostatic parameters such as atomic point charges and dipole moments in ΔAla-containing peptides with internal (conformation) and external (solvent) effects are discussed. © Munksgaard 1995.     

Conformational investigation of α, β-dehydropeptides

The crystal structure of Ac-Pro-ΔVal-NHCH₃ was examined to determine the influence of the α,β-dehydrovaline residue on the nature of peptide conformation. The peptide crystallizes from methanol-diethyl ether solution at 4° in needle-shaped form in orthorhombic space group P2₁2₁2₁ with a= 11.384(2) Å, b = 13.277(2) Å, c = 9.942(1) Å. V = 1502.7(4) ų Z = 4, D_m= 1.17 g cm^?3 and D_c=1.18 g cm^?3 The structure was solved by direct methods using SHELXS-86 and refined to an R value of 0.057 for 1922 observed reflections. The peptide is found to adopt a β-bend between the type I and the type III conformation with φ₁=?68.3(4)°, ψ₁=? 20.1(4)°, φ₂=?73.5(4)°= and Ψ₂=?14.1(4)°=. An intramolecular hydrogen bond between the carbonyl oxygen of ith residue and the NH of (i+ 3)th residue stabilizes the β-bend. An additional intermolecular N.,.O hydrogen bond joins molecules into infinite chains. In the literature described crystal structures of peptides having a single α,β-dehydroamino acid residue in the (i+ 2) position and forming a β-bend reveal a type II conformation.     

Studies on β-turn of peptides

The effect of changing 1st and 4th amino acid residues on β-turn preference of tetrapeptide sequences was studied by use of CD spectra of the chromophoric derivatives, which have Dnp- and pNA-groups as the amino and carboxyl substituents, respectively. The effect was examined with the tetrapeptides having such sequences at the 2nd and 3rd positions as -L-Pro-L-Asn-, -L-Pro-Gly-, -L-Pro-D-Ala-, -L-Ala-D-Leu-, -L-Ala-L-Pro-, and -D-Ala-L-Pro-. The β-turn preferences estimated from the CD intensities of the bands due to exciton interaction were found to depend largely on the configurations of the 1st and 4th amino acid residues. When 1st and 2nd (or 3rd and 4th) residues had the same configuration, decreased intensity of the CD band was observed even if the internal sequence had high β-turn preference. Terminal Gly residues were favorable for the β-turn conformation in many of the tetrapeptide sequences examined.     

Solid phase synthesis of human growth hormone-releasing factor analogs containing a bicyclic β-turn dipeptide

Three analogs derived from the N-terminal 29-residue fragment of human growth hormone-releasing factor (hGRF) which contained a bicyclic β-turn dipeptide (BTD) at 7-8,8-9, and 9-10 positions were synthesized by solid phase methodology to ascertain if the β-turns are important for the biological activity of hGRF and also to show the applicability of the BTD unit to solid phase synthesis. All three analogs were obtained in good yield and purity indicating that the BTD unit can be used in the usual condition of solid phase synthesis. The capacity of these analogs to release growth hormone (GH) was tested in an in vitro bioassay using rat anterior pituitary cells. All three BTD-containing analogs showed the same maximal GH secretion with parallel dose-response curves to that of hGRF(1-29)NH₂, except their relative potencies were very low.     

Conformational analysis of cyclized dipeptide models for specific types of β-bends by two-dimensional nuclear Overhauser spectroscopy*

The solution conformations in DMSO-d₆ of the two cyclized dipeptides, cyclo(l -alanyl-l -alanyl-e -aminocaproyl) and cyclo(l -alanyl-d -alanyl-e -amino-caproyl), have been analyzed by means of the two-dimensional nuclear Overhauser effect (2D-NOE). The preferred conformations for the two compounds have been deduced by comparing proton-proton distances, derived from the 2D-NOE data and relaxation-time measurements, with the corresponding distances in several possible computed low-energy conformations. The predominant conformations are a type III bend and a type II bend, respectively, for the two compounds. These conclusions agree with those deduced earlier on the basis of infrared and Raman spectra and circular dichroism measurements.     

Secondary structure of a dynamic type I’β‐hairpin peptide

Abstract: A spontaneously folding β‐hairpin peptide (Lys‐Lys‐Tyr‐Thr‐Val‐Ser‐Ile‐Asn‐Gly‐Lys‐Lys‐Ile‐Thr‐Val‐Ser‐Ile) and related cyclic (cyclo‐Gly‐Lys‐Tyr‐Ile‐Asn‐Gly‐Lys‐Ile‐Ile‐Asn) and linear (Ser‐Ile‐Asn‐Gly‐Lys) controls were studied to determine the effects of various factors on secondary structure. Secondary structure was evaluated using circular dichroism (CD) and 1D and 2D ¹H nuclear magnetic resonance (NMR). The effects of chemical modifications in the peptide and various solution conditions were investigated to determine their impact on peptide structure. The β‐hairpin peptide displayed a CD minimum at 216 nm and a TOCSY i + 1 ? i + 2 and i + 2 ?i + 3 interaction, confirming the expected structure. Using NMR α‐proton (H) chemical shifts, the extents of folding of the β‐hairpin and linear control were estimated to be 51 and 25% of the cyclic control (pH 4, 37 °C), which was taken to be maximally folded. Substitution of iso‐aspartic acid for Asn reduced the secondary structure dramatically; substitution of aspartic acid for Asn also disrupted the structure. This result suggests that deamidation in unconstrained β‐turns may have adverse effects on secondary structure. N‐terminal acetylation and extreme pH conditions also reduced structure, while the addition of methanol increased structure.     

“Vinylogs” and “Acetylenylogs” of β-Adrenergic Agents

Vinylogous (Groups III and V ) and acetylenologous (Group IV ) analogs of the classical β-adrenergic agents — stimulants and blockers — were prepared in order to evaluate the effect of degree of saturation, position of unsaturation and rigidity of the chain linking the aromatic ring and the amino containing functional group on biological activity. Derivatives from Group III , which represent 4-aryl-3-butenyl-2-ol-amine analogs of Group II , retained β₁-adrenoceptor antagonist activity albeit substantially less potent (50–200-fold) than that possessed by their aryloxy counterparts. Consistent with the SAR for Group II compounds, substitution at position 2 of the aromatic ring yielded the most potent antagonists ( 5a, 5d, 5g ), with K_B's ranging from 73–93 nM while 3,4-dichloro substitution ( 5e ) markedly reduced antagonist potency (K_B = 2,400 nM). Agonist activity was also noted for 5b and 5d , suggesting that these compounds may be best classified as partial agonists. Representatives from Groups IV and V were inactive as antagonists at the β₁-adrenoceptor confirming the importance of the spatial relationship between the hydroxyl and the amino nitrogen.     

Theoretical π-π* absorption and circular dichroic spectra of cyclic dipeptides

The dipole interaction model, treated by the partially dispersive normal mode method, is used to calculate circular dichroic spectra of cyclo(Gly-Gly), cyclo (Ala-Gly), cyclo(Ala-Ala), cyclo(Pro-Gly), cyclo(Pro-Ala), cyclo(Pro-Val), cyclo (Pro-D-Val), and cyclo(Pro-Pro) in the amide π-π* absorption band near 190 nm. Assuming a standard backbone geometry, spectra which are in fair to good agreement wth experiment are obtained for these molecules. The spectra are predicted to be sensitive to conformations of Pro and Val side chains. The effects of dipeptide ring folding on calculated CD spectra are mostly consistent with those found by other workers, except that it is found that a planar ring conformation of cyclo (Ala-Ala) and cyclo (Ala-Gly) gives predicted spectra comparable to experiment. The same model gives theoretical absorption spectra consistent with available experimental data.     

Synthetic and conformational studies on dehydrophenylalanine containing model peptides

Four model dipeptides containing a Z-dehydrophenylalanine residue (Δ^ZPhe) at the C-terminal, Boc-X-Δ^Z Phe-NHMe (X = Ala (1), Gly (2), Pro (3), and Val (4)), have been synthesised and their solution conformations investigated by 270 MHz ¹H n.m.r. and i.r. spectroscopy. N.m.r. studies on these peptides clearly show the presence of intramolecularly hydrogen bonded structures in CHCl₃ solutions while such structures appear to be absent in the corresponding saturated peptides. This conclusion is also supported by i.r. studies. Studies of the nuclear Overhauser effect provided evidence for the occurrence of a significant population of β-turn structures in solvents like CDCl₃ and (CD₃)₂SO. The observed NOES are consistent with a major contribution from Type II β-turn structure in CDCl₃, while in (CD₃)₂SO solutions there is evidence of a partially extended structure also.     

Turn induction by N-aminoproline Comparison of the Gly-Pro-Gly and Glyψ[CO-NH-N]Pro-Gly sequences

The folded structure induced by the N-aminoproline residue (the hydrazino analogue of proline, denoted hPro) in the Boc-Gly¹-hPro²-Gly³-NHiPr hydrazino tripeptide has been characterized in the solid state by X-ray diffraction, and compared to the usual βII-turn structure in the Boc-Gly¹-Pro²-Gly³⁶-NHiPr cognate tripeptide. It is stabilized by a bifurcated hydrogen bond in which (Gly³)NH interacts with both (Gly¹)CO and (hPro²)N^x. This conformation is retained in CH₂Cl₂ and CHC1₃ solutions, and allows an overall folded conformation of the hydrazino tripeptide in which (iPr)NH is hydrogen-bonded to (Boc)CO. The hPro α-hydrazino acid residue appears to promote a local folded structure, and might behave as a β-turn mimic. © Munksgaard 1994.