Proc. Nat. Acad. Sci. USA Vol. 73, No. 4, pp. 1203-1206, April 1976
Biophysics
Theoretical studies on Pro-Leu-Gly-NH2 conformation (three-dimensional structure/potential function calculations/melanotropin release inhibiting factor/peptide)
SUNGZONG KANG* AND RODERICH WALTERt*§ * Department of Pharmacology, Mount Sinai School of Medicine, City University of New York, New York, N.Y. 10029; t Department of Physiology and Biophysics, University of Illinois Medical Center, Chicago, II. 60612; and t The Medical Research Center, Brookhaven National Laboratory Upton, New York
11973
Communicated by Maurice Goldhaber, February 2,1976
Classical potential function calculations ABSTRACT were carried out on the hypothalamic factor Pro-Leu-GlyNH2. The results indicate that the proposed 1O-membered, hydrogen-bonded f-turn conformation of this tripeptide is a strongly preferred structure. Its stability appears to be inherent in the rather rigid backbone conformation of the leucine residue rather than the hydrogen bond between the carboxamide proton of glycinamide and the C=O of the proline moiety; the glycinamide has little influence on the 04 of the leucine backbone structure. The type II Mturn structure of the Pro-Leu-Gly-NH2 is preferred.
Several years ago it was proposed that the hypothalamic factor Pro-Leu-Gly-NH2, enzymatically derived from its prohormone oxytocin, is an inhibiting factor for the release of pituitary melanotropin (melanocyte-stimulating hormone, MSH) (1). This biologically active tripeptide was subsequently isolated from bovine pituitary extracts (2) and it has since been found to possess additional, extrapituitary activities (3, 4). On the basis of 1H nuclear magnetic resonance in dimethylsulfoxide and preliminary crystallographic data, Walter et al. proposed a preferred conformation of Pro-Leu-Gly-NH2, consisting of a 10-membered fl-turn (4 - 1) closed by a hydrogen bond (5). This structure was confirmed by the x-ray crystallographic study of Reed and Johnson (6). Measurements of spin-lattice relaxation times of the carbon-13 nuclei of Pro-Leu-Gly-NH2 by Deslauriers et al. (7) showed this tripeptide to have a compact but flexible structure in aqueous medium. In addition, classical potential function calculations which incorporated electrostatic, torsion, and hydrogen-bond contributions revealed a large number of preferred conformations including the 4 - 1 fl-turn (8). The present conformational energy calculations using the
classical Lennard-Jones potential function, which includes an electrostatic term, represent a further attempt to ascertain whether the stability of the proposed structure of ProLeu-Gly-NH2 is primarily due to the hydrogen bonding energy or is actually inherent in the backbone of this tripeptide. A second goal of this study was to determine whether there was a preference for one of the two theoretically possible types of f-turn conformations (9-12). One is the type I, in which the leucine side chain would occupy an axial position, and the other is the type II with an equatorial position of the leucine side chain. Knowledge of the preferred type
of f-turn of Pro-Leu-Gly-NH2 may be of particular interest when one considers the biological activity of this hypothalamic factor and its analogs. Abbreviations: tg, trans-gauche; gt, gauche-trans. § To whom correspondence should be addressed at the Department of Physiology and Biophysics, University of Illinois at the Medical Center, P.O. Box 6998, Chicago, III. 60680.
1203
METHODS The Lennard-Jones 6-12 potential function was used, including an electrostatic term, and the procedure has been described in detail (13). The net charges in the Coulomb term were calculated by the INDO molecular orbital method. The molecular geometry of Pro-Leu-Gly-NH2 (Fig. 1) has been constructed using the reported x-ray crystallographic data (6) with the naturally occurring Pro(S)-Leu(S) absolute configuration. The description and direction of rotation followed the IUPAC-IUB convention (14) except that the angles were scanned from O to 3600 (see figures). Planar peptide bonds were assumed throughout the calculations. RESULTS AND DISCUSSION Leucine side chain Two preferred leucine side chain conformations were observed; one is the gauche-trans (gt) form where xI[N(Leu)-Ca(Leu)-Cfl-Cy] = 300', and X2[Ca-Cfl-CYCB1] = 3000 and X2' [Ca-Cl-Cy-Cb2] = 180°, the other is the trans-gauche (tg) form where xI = 180° and X2 = 180°, and X2' = 60°. In the gt form the leucine side chain is gauche relative to the peptide nitrogen and trans to the Ca of the leucine backbone. These two preferred leucine side chain conformations are virtually independent of the conformation of the rest of the peptide backbone. The gt conformation has been observed in crystal structures of small leucine-containing peptides. Peptide backbone conformations Examination of molecular models and our calculations indicate that 4q of proline has little influence on other backbone rotations of the molecule and hence conformations of this tripeptide should largely depend on the 4-) of leucine and glycinamide. Instead of performing energy-minimization calculations, we have generated a series of leucine 4-)4 maps at various fixed 0-1p values of glycinamide. For the gt leucine side chain conformation and fixed A3 = 700 and jP3 = 2000 of the glycine backbone (the lowest energy 43-i/3 form) the leucine )242 map shows three energy minima; a global minimum near )2 = 280°, P2 = 1000, and two local minima near 2 = 600, i12 = 1600 and near 42 = 3000 and *2 = 3200 (Fig. 2). The global minimum near 02 = 2800 and V2 = 1000 is elongated along the /2 of leucine (Fig. 2). It is interesting to note that these three energy minima are observed independent of the variation of the glycine 3s-4s although minor energy shifts due to the glycine 4)s-4, variation occur within the designated energy minima. Some of these observations are given in Fig. 3. For the tg conformation of the leucine side chain, the global minimum is again near X2 =
Biophysics: Kang and Walter
1204
Proc. Nat. Acad. Sci. USA 73 (1976) 360 300
240 2
180 FIG. 1. Molecular structure of Pro-Leu-Gly-NH2 for a conformation of 4,i(Pro) = 1530, 412(Leu) = 3000, 412(Leu) = 1200, 43(Gly) = 700, and V&3(Gly) = 400.
3W0 and 4/2 = 1200 and elongated along the 2 angle of the leucine backbone (Fig. 4). The leucine 42-02 map is remark-
ably independent of the /3-1/3 variation of the glycine portion. Some of these observations are given in Fig. 5. As shown in Figs. 3 and 5, the global minimum is elongated along the k2 of leucine for the gt side chain conformation, and along the 42 for the tg conformation. Fig. 6 shows that as the side chain moves from the gt conformation to the tg conformation, the shape of the global minimum changes gradually from p2-elongation to +2-elongation [D(Xl =
-60°)
--
G(XI = -80°) -- B(XI = -160°) -- A(Xj = 1800)
in Fig. 6]. This observation clearly indicates that the global minimum is not shifted to other locations by the conformation of the leucine side chain. Using the classical potential function it appears that the
'2t2 FIG. 3. 024-2 energy map of Pro-Leu-Gly-NH2 for different values of (X3, 4,3). The side chain has a gt conformation. The lines indicate 6 kcal/mol (1 kcal = 4.184 kJ) relative to the lowest energy of the (43 = 700, 4P3 = 2000)-conformation as zero. The (4'3, 43) angles were fixed as follows: - - for (700, 2000), --- for (2600, - for (-1190, 1130), - - for (-570, -470), - -- 1600), - - - - - for (1800, 1800) and - for (700, 300). & indicates the conformation found in the crystal (6). - -
--
over-all conformation of Pro-Leu-Gly-NH2 is mainly determined by the relative rigidity of the backbone of the leucine residue near 42 = 2900 and {2 = 1200. This suggestion is based upon three observations: (i) the global minimum near 02 = 2900 and 5,62 = 1200 is the lowest and widest; (ii) it is independent of the position of the glycine backbone; and (iii) it is independent of the position of the leucine side chain. In agreement with this study, the carbon-1S relaxation data by Deslauriers et al. (7) also revealed a greater ri-
360 300
240 80-
120 60 -e4
0
60
120
180
240
300
360
2
k2-42 energy map of Pro-Leu-Gly-NH2 for fixed values of 03 = 700 and 03 = 2000. Side chain is in the gt conformation. A indicates the conformation found in the crystal (6). FIG. 2.
FIG. 4. 02-4'2 energy map of Pro-Leu-Gly-NH2 for fixed values of 03 = 700 and 4'3 = 2000. Side chain of Leu has a tg conformation. The numbers indicate energy in kcal/mol. & indicates the conformation found in the crystal (6).
Proc. Nat. Acad. ScA. USA 73 (1976)
Biophysics: Kang and Walter
1205
¢2I5LAt 180
120
60
60
0
120
180
240
300
360
120
180 3 FIG. 7. 03k3 energy map of Pro-Leu-Gly-NH2 for fixed values of '12 = 3000 and '12 = 1200. The lowest energy of (X13-+13) is (700, 2000).
60
3/.
'2 FIG. 5. 42-42 energy map of Pro-Leu-Gly-NH2 for different values of (Xk3, *3). The side chain has a tg conformation. The lines indicate 6 kcal/mol relative to the lowest energy of the (43 = 700,
03 = 2000)-conformation as zero. The (03, '13) angles were fixed as - - for (700, 2000), --- for (260°, 1600), - - for - for (-119°, 1130), for (-570, -470), -for (1000, 400). A indicates the conformation (-500, 800), and
follows:
found in the crystal (6).
gidity for the leucine backbone as compared to the glycine moiety. It is noteworthy that the lowest energy conformation of the leucine backbone near 02 = 240-3200 and 4i2 = 80(1400 is the optimum condition for the proposed 10membered #-turn hydrogen-bonded conformation of ProLeu-Gly-NH2 in dimethylsulfoxide (5) and in the crystalline 360 300 240
2
.0
180
120
state (6) even without taking hydrogen bonding energies explicitly into account during the calculations. These results are in agreement with studies on the preferred conformation of the tripeptide backbone structure per se, whether a hydrogen bond function was included (16) or not (V. Sasisekharan, unpublished). The importance of the intrinsic con-
formation of the peptide backbone in determining structural homogeneity of cyclic peptides was pointed out in a series of crystallographic studies of Titlestad, Dale, and collaborators (17). The glycine backbone, on the other hand, is so flexible that the preferred conformation of glycinamide probably depends largely on the presence of hydrogen bonds, the crystal packing, interaction with the solvent or receptors, and other environmental factors (Fig. 7). However, it is observed that at the extended Gly A's torsional angle the lowest k2-1/2 energy is near Gly A3 = 70°. This is again an angle compatible for a 10-membered ,8-turn conformation (5, 6); in the crystal structure an angle of 72° was found. Theoretically a 1O-membered R-turn cyclic structure can assume different configurations (9-12) (types I, I', II, and II'). According to our calculations, the preferred form of Pro-Leu-Gly-NH2 is the type II, with /2(Leu) = 3000 and 4i2(Leu) = 120°, and the leucine side chain in an equatorial position relative to the plane of the 10-membered backbone ring. This is in good agreement with the observed conformation of this molecule in the crystal (6). This work was supported by USPHS Grants MH-17489 and AM18399.
O _
0
1. Celis, M. E., Taleisnik, S. & Walter, R. (1971) Proc. Nat.
60
120
180
240
300
360
Yf2 FIG. 6. 024'2 energy map of Pro-Leu-Gly-NH2 for fixed values of 13 = 700 and = 2000. The side chain of leucine has been varied as follows: A for tg conformation at xi = 1800, X2 = 1800, and X2' = 60°. B for tg conformation at Xi = 1600, X2 = 1800, and X2' = 600. C for gt conformation at X2 = 3000, X2 = 3000, and X2' = 1800. D for gt conformation at XI = 2800, Xi = 1800, and X2' = 3000. indicates the conformation found in the crystal (6). 1)
Acad. Sci. USA 68,1428-1433. 2. Nair, R. M. G., Kastin, A. J. & Schally, A. V. (1971) Biochem. Biophys. Res. Commun. 43, 1376-1381. 3. Plotnikoff, N. P., Kastin, A. J., Anderson, M. S. & Schally, A. V. (1971) Life Sci. 10, 1279-1283. 4. Kastin, A. J., Plotnikoff, N. P., Nair, R. M. G., Redding, T. W. & Anderson, M. S. (1973) in Hypothalamic Hypophysiotropic Hormones: Physiological and Clinical Studies, ed. Gual, C. (Excerpta Medica, Amsterdam), pp. 159ff. 5. Walter, R., Bernal I. & Johnson, L. F. (1972) in Chemistry
1206
6.
7. 8.
9.
Biophysics: Kang and Walter and Biology of Peptides, ed. Meienhofer, J. (Ann Arbor Publ., Ann Arbor, Mich.), pp. 131-135. Reed, L. L. & Johnson, P. L. (1973) J. Am. Chem. Soc. 95, 7523-7524. Deslauriers, R., Walter, R. & Smith, I. C. P. (1973) FEBS Lett. 37,27-32. Ralston, E., De Coen, J.-L. & Walter, R. (1974) Proc. Nat. Acad. Sci. USA 71, 1142-1144. Geddes, A. J., Parker, K. D., Atkins, E. D. T. & Beighton, E.
(1968) J. Mol. Biol. 32, 343-358. 10. Venkatachalam, C. M. (1968) Blopolymers 6, 1425-1436. 11. Urry, D. W. & Ohnishi, M. (1970) in Spectroscopic Approaches to Biomolecular Conformation, ed. Urry, D. W. (American Medical Association, Chicago), pp. 263ff.
Proc. Nat. Acad. Sci. USA 73 (1976) 12. Crawford, J. L., Lipscomb, W. N. & Schellman, C. G. (1973) Proc. Nat. Acad. Sci. USA 70,538-542. 13. Kang, S., Froimowitz, M. & Hankins, D. (1974) J. Theor. Biol.
44,337-347. 14. IUPAC-IUB Commission on Biochemical Nomenclature
(1970) Biochemistry 9,3471-3478. 15. Ueki, T., Ashida, T., Kakudo, M., Sasada, Y. & Katsube, Y. (1969) Acta Crystallogr. Sect. B 25, 1840-1849. 16. Chandrasekaran, R., Lakshiminarayanan, A. V., Pandya, U. V. & Ramachandran, G. N. (1973) Biochim. Biophys. Acta 303, 14-27. 17. Declercq, J. P., Germain, G., van Meerssche, M., Debaerdemaeker, T., Dale, J. & Titlestad, K. (1975) Bull. Soc. Chim.
Beig. 84,275-287.
Biophysics
Theoretical studies on Pro-Leu-Gly-NH2 conformation (three-dimensional structure/potential function calculations/melanotropin release inhibiting factor/peptide)
SUNGZONG KANG* AND RODERICH WALTERt*§ * Department of Pharmacology, Mount Sinai School of Medicine, City University of New York, New York, N.Y. 10029; t Department of Physiology and Biophysics, University of Illinois Medical Center, Chicago, II. 60612; and t The Medical Research Center, Brookhaven National Laboratory Upton, New York
11973
Communicated by Maurice Goldhaber, February 2,1976
Classical potential function calculations ABSTRACT were carried out on the hypothalamic factor Pro-Leu-GlyNH2. The results indicate that the proposed 1O-membered, hydrogen-bonded f-turn conformation of this tripeptide is a strongly preferred structure. Its stability appears to be inherent in the rather rigid backbone conformation of the leucine residue rather than the hydrogen bond between the carboxamide proton of glycinamide and the C=O of the proline moiety; the glycinamide has little influence on the 04 of the leucine backbone structure. The type II Mturn structure of the Pro-Leu-Gly-NH2 is preferred.
Several years ago it was proposed that the hypothalamic factor Pro-Leu-Gly-NH2, enzymatically derived from its prohormone oxytocin, is an inhibiting factor for the release of pituitary melanotropin (melanocyte-stimulating hormone, MSH) (1). This biologically active tripeptide was subsequently isolated from bovine pituitary extracts (2) and it has since been found to possess additional, extrapituitary activities (3, 4). On the basis of 1H nuclear magnetic resonance in dimethylsulfoxide and preliminary crystallographic data, Walter et al. proposed a preferred conformation of Pro-Leu-Gly-NH2, consisting of a 10-membered fl-turn (4 - 1) closed by a hydrogen bond (5). This structure was confirmed by the x-ray crystallographic study of Reed and Johnson (6). Measurements of spin-lattice relaxation times of the carbon-13 nuclei of Pro-Leu-Gly-NH2 by Deslauriers et al. (7) showed this tripeptide to have a compact but flexible structure in aqueous medium. In addition, classical potential function calculations which incorporated electrostatic, torsion, and hydrogen-bond contributions revealed a large number of preferred conformations including the 4 - 1 fl-turn (8). The present conformational energy calculations using the
classical Lennard-Jones potential function, which includes an electrostatic term, represent a further attempt to ascertain whether the stability of the proposed structure of ProLeu-Gly-NH2 is primarily due to the hydrogen bonding energy or is actually inherent in the backbone of this tripeptide. A second goal of this study was to determine whether there was a preference for one of the two theoretically possible types of f-turn conformations (9-12). One is the type I, in which the leucine side chain would occupy an axial position, and the other is the type II with an equatorial position of the leucine side chain. Knowledge of the preferred type
of f-turn of Pro-Leu-Gly-NH2 may be of particular interest when one considers the biological activity of this hypothalamic factor and its analogs. Abbreviations: tg, trans-gauche; gt, gauche-trans. § To whom correspondence should be addressed at the Department of Physiology and Biophysics, University of Illinois at the Medical Center, P.O. Box 6998, Chicago, III. 60680.
1203
METHODS The Lennard-Jones 6-12 potential function was used, including an electrostatic term, and the procedure has been described in detail (13). The net charges in the Coulomb term were calculated by the INDO molecular orbital method. The molecular geometry of Pro-Leu-Gly-NH2 (Fig. 1) has been constructed using the reported x-ray crystallographic data (6) with the naturally occurring Pro(S)-Leu(S) absolute configuration. The description and direction of rotation followed the IUPAC-IUB convention (14) except that the angles were scanned from O to 3600 (see figures). Planar peptide bonds were assumed throughout the calculations. RESULTS AND DISCUSSION Leucine side chain Two preferred leucine side chain conformations were observed; one is the gauche-trans (gt) form where xI[N(Leu)-Ca(Leu)-Cfl-Cy] = 300', and X2[Ca-Cfl-CYCB1] = 3000 and X2' [Ca-Cl-Cy-Cb2] = 180°, the other is the trans-gauche (tg) form where xI = 180° and X2 = 180°, and X2' = 60°. In the gt form the leucine side chain is gauche relative to the peptide nitrogen and trans to the Ca of the leucine backbone. These two preferred leucine side chain conformations are virtually independent of the conformation of the rest of the peptide backbone. The gt conformation has been observed in crystal structures of small leucine-containing peptides. Peptide backbone conformations Examination of molecular models and our calculations indicate that 4q of proline has little influence on other backbone rotations of the molecule and hence conformations of this tripeptide should largely depend on the 4-) of leucine and glycinamide. Instead of performing energy-minimization calculations, we have generated a series of leucine 4-)4 maps at various fixed 0-1p values of glycinamide. For the gt leucine side chain conformation and fixed A3 = 700 and jP3 = 2000 of the glycine backbone (the lowest energy 43-i/3 form) the leucine )242 map shows three energy minima; a global minimum near )2 = 280°, P2 = 1000, and two local minima near 2 = 600, i12 = 1600 and near 42 = 3000 and *2 = 3200 (Fig. 2). The global minimum near 02 = 2800 and V2 = 1000 is elongated along the /2 of leucine (Fig. 2). It is interesting to note that these three energy minima are observed independent of the variation of the glycine 3s-4s although minor energy shifts due to the glycine 4)s-4, variation occur within the designated energy minima. Some of these observations are given in Fig. 3. For the tg conformation of the leucine side chain, the global minimum is again near X2 =
Biophysics: Kang and Walter
1204
Proc. Nat. Acad. Sci. USA 73 (1976) 360 300
240 2
180 FIG. 1. Molecular structure of Pro-Leu-Gly-NH2 for a conformation of 4,i(Pro) = 1530, 412(Leu) = 3000, 412(Leu) = 1200, 43(Gly) = 700, and V&3(Gly) = 400.
3W0 and 4/2 = 1200 and elongated along the 2 angle of the leucine backbone (Fig. 4). The leucine 42-02 map is remark-
ably independent of the /3-1/3 variation of the glycine portion. Some of these observations are given in Fig. 5. As shown in Figs. 3 and 5, the global minimum is elongated along the k2 of leucine for the gt side chain conformation, and along the 42 for the tg conformation. Fig. 6 shows that as the side chain moves from the gt conformation to the tg conformation, the shape of the global minimum changes gradually from p2-elongation to +2-elongation [D(Xl =
-60°)
--
G(XI = -80°) -- B(XI = -160°) -- A(Xj = 1800)
in Fig. 6]. This observation clearly indicates that the global minimum is not shifted to other locations by the conformation of the leucine side chain. Using the classical potential function it appears that the
'2t2 FIG. 3. 024-2 energy map of Pro-Leu-Gly-NH2 for different values of (X3, 4,3). The side chain has a gt conformation. The lines indicate 6 kcal/mol (1 kcal = 4.184 kJ) relative to the lowest energy of the (43 = 700, 4P3 = 2000)-conformation as zero. The (4'3, 43) angles were fixed as follows: - - for (700, 2000), --- for (2600, - for (-1190, 1130), - - for (-570, -470), - -- 1600), - - - - - for (1800, 1800) and - for (700, 300). & indicates the conformation found in the crystal (6). - -
--
over-all conformation of Pro-Leu-Gly-NH2 is mainly determined by the relative rigidity of the backbone of the leucine residue near 42 = 2900 and {2 = 1200. This suggestion is based upon three observations: (i) the global minimum near 02 = 2900 and 5,62 = 1200 is the lowest and widest; (ii) it is independent of the position of the glycine backbone; and (iii) it is independent of the position of the leucine side chain. In agreement with this study, the carbon-1S relaxation data by Deslauriers et al. (7) also revealed a greater ri-
360 300
240 80-
120 60 -e4
0
60
120
180
240
300
360
2
k2-42 energy map of Pro-Leu-Gly-NH2 for fixed values of 03 = 700 and 03 = 2000. Side chain is in the gt conformation. A indicates the conformation found in the crystal (6). FIG. 2.
FIG. 4. 02-4'2 energy map of Pro-Leu-Gly-NH2 for fixed values of 03 = 700 and 4'3 = 2000. Side chain of Leu has a tg conformation. The numbers indicate energy in kcal/mol. & indicates the conformation found in the crystal (6).
Proc. Nat. Acad. ScA. USA 73 (1976)
Biophysics: Kang and Walter
1205
¢2I5LAt 180
120
60
60
0
120
180
240
300
360
120
180 3 FIG. 7. 03k3 energy map of Pro-Leu-Gly-NH2 for fixed values of '12 = 3000 and '12 = 1200. The lowest energy of (X13-+13) is (700, 2000).
60
3/.
'2 FIG. 5. 42-42 energy map of Pro-Leu-Gly-NH2 for different values of (Xk3, *3). The side chain has a tg conformation. The lines indicate 6 kcal/mol relative to the lowest energy of the (43 = 700,
03 = 2000)-conformation as zero. The (03, '13) angles were fixed as - - for (700, 2000), --- for (260°, 1600), - - for - for (-119°, 1130), for (-570, -470), -for (1000, 400). A indicates the conformation (-500, 800), and
follows:
found in the crystal (6).
gidity for the leucine backbone as compared to the glycine moiety. It is noteworthy that the lowest energy conformation of the leucine backbone near 02 = 240-3200 and 4i2 = 80(1400 is the optimum condition for the proposed 10membered #-turn hydrogen-bonded conformation of ProLeu-Gly-NH2 in dimethylsulfoxide (5) and in the crystalline 360 300 240
2
.0
180
120
state (6) even without taking hydrogen bonding energies explicitly into account during the calculations. These results are in agreement with studies on the preferred conformation of the tripeptide backbone structure per se, whether a hydrogen bond function was included (16) or not (V. Sasisekharan, unpublished). The importance of the intrinsic con-
formation of the peptide backbone in determining structural homogeneity of cyclic peptides was pointed out in a series of crystallographic studies of Titlestad, Dale, and collaborators (17). The glycine backbone, on the other hand, is so flexible that the preferred conformation of glycinamide probably depends largely on the presence of hydrogen bonds, the crystal packing, interaction with the solvent or receptors, and other environmental factors (Fig. 7). However, it is observed that at the extended Gly A's torsional angle the lowest k2-1/2 energy is near Gly A3 = 70°. This is again an angle compatible for a 10-membered ,8-turn conformation (5, 6); in the crystal structure an angle of 72° was found. Theoretically a 1O-membered R-turn cyclic structure can assume different configurations (9-12) (types I, I', II, and II'). According to our calculations, the preferred form of Pro-Leu-Gly-NH2 is the type II, with /2(Leu) = 3000 and 4i2(Leu) = 120°, and the leucine side chain in an equatorial position relative to the plane of the 10-membered backbone ring. This is in good agreement with the observed conformation of this molecule in the crystal (6). This work was supported by USPHS Grants MH-17489 and AM18399.
O _
0
1. Celis, M. E., Taleisnik, S. & Walter, R. (1971) Proc. Nat.
60
120
180
240
300
360
Yf2 FIG. 6. 024'2 energy map of Pro-Leu-Gly-NH2 for fixed values of 13 = 700 and = 2000. The side chain of leucine has been varied as follows: A for tg conformation at xi = 1800, X2 = 1800, and X2' = 60°. B for tg conformation at Xi = 1600, X2 = 1800, and X2' = 600. C for gt conformation at X2 = 3000, X2 = 3000, and X2' = 1800. D for gt conformation at XI = 2800, Xi = 1800, and X2' = 3000. indicates the conformation found in the crystal (6). 1)
Acad. Sci. USA 68,1428-1433. 2. Nair, R. M. G., Kastin, A. J. & Schally, A. V. (1971) Biochem. Biophys. Res. Commun. 43, 1376-1381. 3. Plotnikoff, N. P., Kastin, A. J., Anderson, M. S. & Schally, A. V. (1971) Life Sci. 10, 1279-1283. 4. Kastin, A. J., Plotnikoff, N. P., Nair, R. M. G., Redding, T. W. & Anderson, M. S. (1973) in Hypothalamic Hypophysiotropic Hormones: Physiological and Clinical Studies, ed. Gual, C. (Excerpta Medica, Amsterdam), pp. 159ff. 5. Walter, R., Bernal I. & Johnson, L. F. (1972) in Chemistry
1206
6.
7. 8.
9.
Biophysics: Kang and Walter and Biology of Peptides, ed. Meienhofer, J. (Ann Arbor Publ., Ann Arbor, Mich.), pp. 131-135. Reed, L. L. & Johnson, P. L. (1973) J. Am. Chem. Soc. 95, 7523-7524. Deslauriers, R., Walter, R. & Smith, I. C. P. (1973) FEBS Lett. 37,27-32. Ralston, E., De Coen, J.-L. & Walter, R. (1974) Proc. Nat. Acad. Sci. USA 71, 1142-1144. Geddes, A. J., Parker, K. D., Atkins, E. D. T. & Beighton, E.
(1968) J. Mol. Biol. 32, 343-358. 10. Venkatachalam, C. M. (1968) Blopolymers 6, 1425-1436. 11. Urry, D. W. & Ohnishi, M. (1970) in Spectroscopic Approaches to Biomolecular Conformation, ed. Urry, D. W. (American Medical Association, Chicago), pp. 263ff.
Proc. Nat. Acad. Sci. USA 73 (1976) 12. Crawford, J. L., Lipscomb, W. N. & Schellman, C. G. (1973) Proc. Nat. Acad. Sci. USA 70,538-542. 13. Kang, S., Froimowitz, M. & Hankins, D. (1974) J. Theor. Biol.
44,337-347. 14. IUPAC-IUB Commission on Biochemical Nomenclature
(1970) Biochemistry 9,3471-3478. 15. Ueki, T., Ashida, T., Kakudo, M., Sasada, Y. & Katsube, Y. (1969) Acta Crystallogr. Sect. B 25, 1840-1849. 16. Chandrasekaran, R., Lakshiminarayanan, A. V., Pandya, U. V. & Ramachandran, G. N. (1973) Biochim. Biophys. Acta 303, 14-27. 17. Declercq, J. P., Germain, G., van Meerssche, M., Debaerdemaeker, T., Dale, J. & Titlestad, K. (1975) Bull. Soc. Chim.
Beig. 84,275-287.
