Preferred Conformation of Peptides from CataSymmetrically Disubstituted Glycines: Aromatic Residues M. CRISMA,' G. VANE,' G. M. BONORA,' C. T O N I O L O , ' F. LEY,' V. BARONE,3 F. FRATERNAL1,3 P. M. HARDY,' and H. 1. S. M A I A '
' Biopolymer Research Center, C N R , Department of Organic Chemistry, University of Padova, 351 31 Padova, 'Institute of Chemistry, University of Basilicata, 85100 Potenza, 'Department of Chemistry, University of Naples, 801 34 Naples, 4Department of Chemistry, University of Exeter, Exeter EX4 4QD, Devon, England, 5Center tor Pure and Applied Chemistry, University of Minho, 4700 Braga, Portugal
SYNOPSIS
The conformational preference of C "*"-diphenylglycine ( D@g) and C"*"-dibenzylglycine (Dbz) residues was assessed in selected derivatives and small peptides by conformational energy computations, ir absorption, 'H-nmr, and x-ray diffraction. Conformational energy computations on the two monopeptides strongly support the view that these C "*"-symmetrically disubstituted glycines are conformationally restricted and that their minimum energy conformation falls in the fully extended (C,) region. The results of the theoretical analyses appear to be in agreement with the solution and crystal-state structural propensities of three derivatives and seven di- and tripeptides.
INTRODUCTION
tides containing either a C "*"-diphenylglycine (Ddg ) or a C "*"-dibenzylglycine (Dbz ) residue.
The C ","-symmetricallydisubstituted glycyl residues can be exploited to modify the conformation of the peptide backbone of bioactive peptides in a very specific way. The set of d,$ torsion angles depends upon the size of the side chains of the incorporated residue. In this connection recent studies of model pept,ides have suggested that C "*"-dimethylglycine ( cu-aminoisobutyric acid, Aib) and l-aminocycloalkane carboxylic acid residues, Ac,c (Figure 1) strongly prefer folded backbone conformations in the 31,,/cu-helical region of the $,$ space ( 6 = +60 k 20°, $ = t 3 0 -t 20"); by contrast, C","-diethylglycine ( Deg) and C"3"-di-n-propylglycine (Dpg) residues, with acyclic side chains longer than methyl groups, preferentially adopt the fully extended (C,) conformation ($,$ s 180°, 180'; for recent review articles, see Refs. 1-4). As a part of a program aimed at investigating the preferred conformations of C ""'-symmetrically disubstituted glycyl residues, we report here the structural characterization (by using conformational energy computations, ir absorption, 'H-nmr, and xray diffraction) of a variety of derivatives and pepHiopolymers, Vol. :31, 637-641 (1991) t.. 1991 J o h n Wiley & Sons, Inc.
RESULTS
Table I gives the conformational energy computation data for Ddg and Dbz monopeptides. In both cases the following is relevant: ( a ) The conformational space available is strongly reduced with respect to C"-monosubstituted a-amino acids. ( b ) The most stable conformation is the fully extended ( C5) structure ( I ) . ( c ) Higher energy minima occur in the C7 (11) and 0 1 / 3 , ~(111) helical regions. The ir absorption spectra in a solvent of low polarity, CDC13 (for three examples, see Figure 2 ) are characterized by a concentration-independent band a t 3405-3385 cmpl, where weakly intramolecularly H-bonded N-H stretching vibrators of the C5 type are usually seen. In the 'H-nmr spectra, all Ddg and Dbz NH protons are essentially concentration independent and exhibit remarkably small variations as the perturbing agents DMSO and TEMPO are added t o the CDCl, solution (for two examples, see Figures 3 and 4). Conversely, all other NH protons in the molecules are much more sensitive to DMSO and ccc o o o f i - ~ ~ z ~ / ~ i / o s o s ~ 7 - o 5 ~ o 4 . oTEMPO. o On the basis of these data and on the po637
638
CRISMA ET AL.
H,C
CH3 I -NH-CCO -
-(CH
\ C/ -NH-CO-
I
) Zn-2
CHJ
I
Ac ,c
Aib
w
u
z 4
m
a
0
m m
U
I - NH-C-CO-
- N H - C-CO-
3500
3450
3400
3350
WAVE NUMBE R 0.0
D bz
Figure 1. The C "+-symmetrically disubstituted glycyl residues discussed in this work.
sition (N-terminal) of the Ddg and Dbz residues in the chain of most of the peptides examined we are inclined to conclude that in CDC13 solution the incorporation of these C "'"-symmetrically disubstituted glycines tend to favor the onset of an intramolecularly H-bonded C5 conformation. According to our x-ray diffraction analysis, the three Ddg and the five Dbz derivatives and peptides examined adopt an intramolecularly H-bonded fully extended C5-ring structure in the crystal state (Table 11). Figures 5 and 6 illustrate the molecular structure of two relevant examples. The N 0 intramolecular distances, ranging from 2.520 ( 5) to 2.651 (8) 8, are typical for a C5 conformation. Further evidence for the presence of the strain introduced by the C5 structure is provided by the values of the critical intraring N-C"-C' ( 7 )bond angle [from 101.8(4)" to 105.6(7)", as expected for a pentagonal ring], remarkably compressed with respect to the tetrahedral value. The C5conformation of mC1Ac-Ddg-OH and the three Tfa-protected Dbz dipeptides is additionally stabilized by an intramolecular N-H * X ( X = C1 or F ) interaction. Further, although indirect, support for the occurrence of the intramolecularly H-bonded conformation is given by the observation that the pertinent N-H and C=O groups are in general not involved in the intermolecular H-bonding schemes.
3300
3; 50
(cm -1)
Figure 2. Infrared absorption spectra in the N-H stretching region of Z-D@g-OtBu( A ) , pBrBz-D@g-GlyOMe ( B ) , and pBrBz-D@g-Gly-OtBu( C ) in CDCl, solution. Concentration IO-~M.
strongly support the view that the Ddg and Dbz residues, with C "3"-symmetricallydisubstituted bulky side chains, favor the fully extended, intramolecularly H-bonded C5-ringstructure, in the ( 180", 180") region of the ($,$) conformational map. The agreement between the preferred conformation calculated
--
--
CONCLUSIONS The results of the present theoretical and experimental ( solution and crystal-state ) investigation
'I* OM!%
in C D C l j
' A TEMW in COClj
Figure 3. ( A ) Plot of NH chemical shifts in the 'Hnmr spectra of pBrBz-D@g-Gly-OtBuvs increasing percentages of DMSO in CDC1, solution ( v / v ) . ( B ) Plot of the bandwidth of the NH protons of the same peptide vs increasing percentages of TEMPO ( w / v ) in CDC1, solution. Concentration 1.3 X 10-3M.
PREFERRED CONFORMATION OF PEPTIDES
Table I
6,$ Backbone Torsion Angles Conformation
Ac-Ddg-NHMe
I I1 I11
a
639
(") and Energies (kcal/mol) for Ac-X-NHMe (X = D6g and Dbz)"
dJ
+
176.3 -75.8 -56.2
176.6 72.2 -46.4
m 0.0 1.9 4.7
Conformation
I
Ac-Dbz-NHMe
I1
I11
dJ
4
AE
180.0 -59.4 -60.0
180.0 102.6 -33.0
0.0 8.3 4.0
Abbreviations used: Ac, acetyl; NHMe, methylamino.
Table I1 T Bond Angle (") and 4, J. Backbone Torsion Angles (") for the D$g and Dbz Residues from the X-Ray Diffraction Analyses of Their Derivatives and Peptides
mC1Ac-DdJg-OH Z-DGg-OtBu pBrBz-DdJg-Gly-OMe
102.1(3) 103.9(2) 101.8(4)
177.8(3) 175.6(3) 155.5(6)
174.7(3) 178.4(3) -146.4(5)
mC1Ac-Dbz-OH Tfa-Dbz-Gly-DBH Tfa-Dbz-L-Phe-OMe Tfa-Dbz-L-Phe-OtBu Z-Glv-Dbz-Glv-OH
104.6(4) 104.8(2) 103.5(4) 103.4(3) 105.6(7)
-179.2(5) 177.9(4) 179.7(4) 175.9(4) -178.9(8)
-173.6(4) 176.4(4) 178.2(4) 180.0(4) 179.3(7)
a Abbreviations u s e d mCIAc, monochloroacetyl; Z, benzyloxycarbonyl; pBrBz, para-bromobenzoyl; Tfa, trifluoroacetyl; OtBu, tertbutoxy; OMe, methoxy; DBH, N,N'-dibenzylacylhydrazido.
in vacuo and that found in the crystal state and in a solvent of low polarity highlights a remarkable stability for this structure. Therefore, the incorporation of these residues into bioactive linear peptides
*I*OMSO h CDCL3
might result in the marked stabilization of the fully extended conformation. In previous studies from this laboratory it has been shown that the most populated conformation
*A TEMPO
in
COCL3
Figure 4. ( A ) Plot of NH chemical shifts in the 'H-nmr spectra of Z-Gly-Dbz-Gly-OtBu us. increasing percentages of DMSO in CDCI3 solution ( v / v ) . ( B ) Plot of the bandwidth of the NH protons of the same peptide us. increasing percentages of TEMPO ( w / v ) in CDC13 solution. Concentration 1.1 X 10-3M.
640
CRISMA ET AL.
Figure 5. Molecular structure of [email protected] intramolecular H-bond is shown as a dashed line.
Figure 6. Molecular structure of Z-Gly-Dbz-Gly-OH. The intramolecular H-bond is shown as a dashed line.
PREFERRED CONFORMATION OF PEPTIDES
for the Deg and Dpg residues is the C5conformation; by contrast, Aib and Ac,c residues strongly prefer the 310/a-helical Therefore, this study confirms that the fully extended conformation becomes more stable than the helical structures when the two side-chain C Batoms are symmetrically substituted but not interconnected in a cyclic system, and in addition, it indicates that this conformational preference should not be related to side-chain aromaticity.
REFERENCES
2. Toniolo, C. & Benedetti, E. ( 1988) ZSZ Atlas Science Biochem. 1, 225-230. 3. Toniolo, C. (1989) Biopolymers 28, 247-257. 4. Toniolo, C. (1990) Int. J . Pept. Protein Res. 35, 287300. 5. Crisma, M., Valle G., Bonora, G. M., De Menego, E., Toniolo, C., Lelj, F., Barone, V. & Fraternali, F. ( 1990) Biopolymers 30, 1-11. 6. Valle, G., Crisma, M., Bonora, G. M., Toniolo, C., Lelj, F., Barone, V., Fraternali, F., Hardy, P. M., LangranGoldsmith, A. & Maia, H. L. S. (1990) J . Chem. SOC. Perkin Trans. 2, 1481-1487.
1. Aubry, A., Boussard, G., Cung, M. T., Marraud, M. &
Vitoux, B. (1988) J . Chim. Phys. Physicochim. Biol. 85, 345-359.
641
Received July 18, 1990 Accepted September 18, 1990
' Biopolymer Research Center, C N R , Department of Organic Chemistry, University of Padova, 351 31 Padova, 'Institute of Chemistry, University of Basilicata, 85100 Potenza, 'Department of Chemistry, University of Naples, 801 34 Naples, 4Department of Chemistry, University of Exeter, Exeter EX4 4QD, Devon, England, 5Center tor Pure and Applied Chemistry, University of Minho, 4700 Braga, Portugal
SYNOPSIS
The conformational preference of C "*"-diphenylglycine ( D@g) and C"*"-dibenzylglycine (Dbz) residues was assessed in selected derivatives and small peptides by conformational energy computations, ir absorption, 'H-nmr, and x-ray diffraction. Conformational energy computations on the two monopeptides strongly support the view that these C "*"-symmetrically disubstituted glycines are conformationally restricted and that their minimum energy conformation falls in the fully extended (C,) region. The results of the theoretical analyses appear to be in agreement with the solution and crystal-state structural propensities of three derivatives and seven di- and tripeptides.
INTRODUCTION
tides containing either a C "*"-diphenylglycine (Ddg ) or a C "*"-dibenzylglycine (Dbz ) residue.
The C ","-symmetricallydisubstituted glycyl residues can be exploited to modify the conformation of the peptide backbone of bioactive peptides in a very specific way. The set of d,$ torsion angles depends upon the size of the side chains of the incorporated residue. In this connection recent studies of model pept,ides have suggested that C "*"-dimethylglycine ( cu-aminoisobutyric acid, Aib) and l-aminocycloalkane carboxylic acid residues, Ac,c (Figure 1) strongly prefer folded backbone conformations in the 31,,/cu-helical region of the $,$ space ( 6 = +60 k 20°, $ = t 3 0 -t 20"); by contrast, C","-diethylglycine ( Deg) and C"3"-di-n-propylglycine (Dpg) residues, with acyclic side chains longer than methyl groups, preferentially adopt the fully extended (C,) conformation ($,$ s 180°, 180'; for recent review articles, see Refs. 1-4). As a part of a program aimed at investigating the preferred conformations of C ""'-symmetrically disubstituted glycyl residues, we report here the structural characterization (by using conformational energy computations, ir absorption, 'H-nmr, and xray diffraction) of a variety of derivatives and pepHiopolymers, Vol. :31, 637-641 (1991) t.. 1991 J o h n Wiley & Sons, Inc.
RESULTS
Table I gives the conformational energy computation data for Ddg and Dbz monopeptides. In both cases the following is relevant: ( a ) The conformational space available is strongly reduced with respect to C"-monosubstituted a-amino acids. ( b ) The most stable conformation is the fully extended ( C5) structure ( I ) . ( c ) Higher energy minima occur in the C7 (11) and 0 1 / 3 , ~(111) helical regions. The ir absorption spectra in a solvent of low polarity, CDC13 (for three examples, see Figure 2 ) are characterized by a concentration-independent band a t 3405-3385 cmpl, where weakly intramolecularly H-bonded N-H stretching vibrators of the C5 type are usually seen. In the 'H-nmr spectra, all Ddg and Dbz NH protons are essentially concentration independent and exhibit remarkably small variations as the perturbing agents DMSO and TEMPO are added t o the CDCl, solution (for two examples, see Figures 3 and 4). Conversely, all other NH protons in the molecules are much more sensitive to DMSO and ccc o o o f i - ~ ~ z ~ / ~ i / o s o s ~ 7 - o 5 ~ o 4 . oTEMPO. o On the basis of these data and on the po637
638
CRISMA ET AL.
H,C
CH3 I -NH-CCO -
-(CH
\ C/ -NH-CO-
I
) Zn-2
CHJ
I
Ac ,c
Aib
w
u
z 4
m
a
0
m m
U
I - NH-C-CO-
- N H - C-CO-
3500
3450
3400
3350
WAVE NUMBE R 0.0
D bz
Figure 1. The C "+-symmetrically disubstituted glycyl residues discussed in this work.
sition (N-terminal) of the Ddg and Dbz residues in the chain of most of the peptides examined we are inclined to conclude that in CDC13 solution the incorporation of these C "'"-symmetrically disubstituted glycines tend to favor the onset of an intramolecularly H-bonded C5 conformation. According to our x-ray diffraction analysis, the three Ddg and the five Dbz derivatives and peptides examined adopt an intramolecularly H-bonded fully extended C5-ring structure in the crystal state (Table 11). Figures 5 and 6 illustrate the molecular structure of two relevant examples. The N 0 intramolecular distances, ranging from 2.520 ( 5) to 2.651 (8) 8, are typical for a C5 conformation. Further evidence for the presence of the strain introduced by the C5 structure is provided by the values of the critical intraring N-C"-C' ( 7 )bond angle [from 101.8(4)" to 105.6(7)", as expected for a pentagonal ring], remarkably compressed with respect to the tetrahedral value. The C5conformation of mC1Ac-Ddg-OH and the three Tfa-protected Dbz dipeptides is additionally stabilized by an intramolecular N-H * X ( X = C1 or F ) interaction. Further, although indirect, support for the occurrence of the intramolecularly H-bonded conformation is given by the observation that the pertinent N-H and C=O groups are in general not involved in the intermolecular H-bonding schemes.
3300
3; 50
(cm -1)
Figure 2. Infrared absorption spectra in the N-H stretching region of Z-D@g-OtBu( A ) , pBrBz-D@g-GlyOMe ( B ) , and pBrBz-D@g-Gly-OtBu( C ) in CDCl, solution. Concentration IO-~M.
strongly support the view that the Ddg and Dbz residues, with C "3"-symmetricallydisubstituted bulky side chains, favor the fully extended, intramolecularly H-bonded C5-ringstructure, in the ( 180", 180") region of the ($,$) conformational map. The agreement between the preferred conformation calculated
--
--
CONCLUSIONS The results of the present theoretical and experimental ( solution and crystal-state ) investigation
'I* OM!%
in C D C l j
' A TEMW in COClj
Figure 3. ( A ) Plot of NH chemical shifts in the 'Hnmr spectra of pBrBz-D@g-Gly-OtBuvs increasing percentages of DMSO in CDC1, solution ( v / v ) . ( B ) Plot of the bandwidth of the NH protons of the same peptide vs increasing percentages of TEMPO ( w / v ) in CDC1, solution. Concentration 1.3 X 10-3M.
PREFERRED CONFORMATION OF PEPTIDES
Table I
6,$ Backbone Torsion Angles Conformation
Ac-Ddg-NHMe
I I1 I11
a
639
(") and Energies (kcal/mol) for Ac-X-NHMe (X = D6g and Dbz)"
dJ
+
176.3 -75.8 -56.2
176.6 72.2 -46.4
m 0.0 1.9 4.7
Conformation
I
Ac-Dbz-NHMe
I1
I11
dJ
4
AE
180.0 -59.4 -60.0
180.0 102.6 -33.0
0.0 8.3 4.0
Abbreviations used: Ac, acetyl; NHMe, methylamino.
Table I1 T Bond Angle (") and 4, J. Backbone Torsion Angles (") for the D$g and Dbz Residues from the X-Ray Diffraction Analyses of Their Derivatives and Peptides
mC1Ac-DdJg-OH Z-DGg-OtBu pBrBz-DdJg-Gly-OMe
102.1(3) 103.9(2) 101.8(4)
177.8(3) 175.6(3) 155.5(6)
174.7(3) 178.4(3) -146.4(5)
mC1Ac-Dbz-OH Tfa-Dbz-Gly-DBH Tfa-Dbz-L-Phe-OMe Tfa-Dbz-L-Phe-OtBu Z-Glv-Dbz-Glv-OH
104.6(4) 104.8(2) 103.5(4) 103.4(3) 105.6(7)
-179.2(5) 177.9(4) 179.7(4) 175.9(4) -178.9(8)
-173.6(4) 176.4(4) 178.2(4) 180.0(4) 179.3(7)
a Abbreviations u s e d mCIAc, monochloroacetyl; Z, benzyloxycarbonyl; pBrBz, para-bromobenzoyl; Tfa, trifluoroacetyl; OtBu, tertbutoxy; OMe, methoxy; DBH, N,N'-dibenzylacylhydrazido.
in vacuo and that found in the crystal state and in a solvent of low polarity highlights a remarkable stability for this structure. Therefore, the incorporation of these residues into bioactive linear peptides
*I*OMSO h CDCL3
might result in the marked stabilization of the fully extended conformation. In previous studies from this laboratory it has been shown that the most populated conformation
*A TEMPO
in
COCL3
Figure 4. ( A ) Plot of NH chemical shifts in the 'H-nmr spectra of Z-Gly-Dbz-Gly-OtBu us. increasing percentages of DMSO in CDCI3 solution ( v / v ) . ( B ) Plot of the bandwidth of the NH protons of the same peptide us. increasing percentages of TEMPO ( w / v ) in CDC13 solution. Concentration 1.1 X 10-3M.
640
CRISMA ET AL.
Figure 5. Molecular structure of [email protected] intramolecular H-bond is shown as a dashed line.
Figure 6. Molecular structure of Z-Gly-Dbz-Gly-OH. The intramolecular H-bond is shown as a dashed line.
PREFERRED CONFORMATION OF PEPTIDES
for the Deg and Dpg residues is the C5conformation; by contrast, Aib and Ac,c residues strongly prefer the 310/a-helical Therefore, this study confirms that the fully extended conformation becomes more stable than the helical structures when the two side-chain C Batoms are symmetrically substituted but not interconnected in a cyclic system, and in addition, it indicates that this conformational preference should not be related to side-chain aromaticity.
REFERENCES
2. Toniolo, C. & Benedetti, E. ( 1988) ZSZ Atlas Science Biochem. 1, 225-230. 3. Toniolo, C. (1989) Biopolymers 28, 247-257. 4. Toniolo, C. (1990) Int. J . Pept. Protein Res. 35, 287300. 5. Crisma, M., Valle G., Bonora, G. M., De Menego, E., Toniolo, C., Lelj, F., Barone, V. & Fraternali, F. ( 1990) Biopolymers 30, 1-11. 6. Valle, G., Crisma, M., Bonora, G. M., Toniolo, C., Lelj, F., Barone, V., Fraternali, F., Hardy, P. M., LangranGoldsmith, A. & Maia, H. L. S. (1990) J . Chem. SOC. Perkin Trans. 2, 1481-1487.
1. Aubry, A., Boussard, G., Cung, M. T., Marraud, M. &
Vitoux, B. (1988) J . Chim. Phys. Physicochim. Biol. 85, 345-359.
641
Received July 18, 1990 Accepted September 18, 1990
