Molecular Dynamics Simulation in Vacuo and in Solution of Cyclolinopeptide A: A Conformational Study MICHELE SAVIANO,* MISAKO AIDA,’ and GlORGlNA CORONGIU I B M Corporation, Scientific Engineering Computations, Department 4 8 B / M S 4 2 8 , Kingston, New York 1 2 4 0 1

SYNOPSIS

The conformation of cyclolinopeptide A [ c- ( Pro-Pro-Phe-Phe-Leu-Ile-Ile-Leu-Val) 1, a naturally occurring peptide with remarkable cytoprotective activity, has been investigated by means of molecular dynamics simulations in various molecular environments. Structural and dynamical properties have been analyzed and compared with those experimentally determined. A detailed analysis of hydrogen bonds is reported.

INTRODUCTION Studies on cyclolinopeptide A ( C L A ) , a cyclic nonapeptide isolated from linseed, have demonstrated that this substance presents remarkable cytoprotective ability.* In particular, it is able t o inhibit the uptake of cholate by hepatocytes. It has been shown that this biological activity is linked to well-defined sequential features shared by another natural cyclic peptide, i.e., antamanide3and by several analogue^.^ The various experimental studies of CLA4 concur in the assumption that the biological activity is critically dependent upon the sequence and conformation of the peptide. Therefore it seems interesting to investigate the conformation-activity relationship of this cyclic peptide from a computational view point. Although in the past many experimental attempts have been made aiming a t resolving the structure of CLA, only recently have a n x-ray structure and a complete nmr study been reported (Figure la).5-7 The study of CLA in solution presents great difficulties, unexpected for small cyclic peptides whose structures are thought to be restricted to a small part of the conformational space. T h e experimental resultsx-” indicate that CLA exists in solution as a

mixture of several conformations; many explanations of this subject have been The room temperature ’H spectra in polar solvents’ were characterized by very broad backbone proton resonances. The strategy of performing nmr experiments a t the lowest possible temperature to “freeze” a single conformation state in a polar solvent was also applied,’ but line broadening was still present. A possible explanation of the existence of multiple conformers even a t very low temperatures is that atoms of CLA form several different hydrogen bonds with solvent molecules. The formation of these hydrogen bonds is also favored by the intrinsic flexibility linked t o the cis-trans isomerism of the two Xxx-Pro bonds. A unique conformational state was proposed’ to be present only in chloroform at low temperature (214 K ) with a structure closely resembling the one from x-ray analysis; both crystalline and solution structures showed a cis peptide bond between the two prolines. In this work we present a comparative analysis of molecular dynamics simulations for CLA in vacuo and in solution, both for a n isolated molecule and for a periodic crystal. We attempt to clarify the characteristic of CLA and explain the experimental observations.

Hiopolyrnws, Vol 31. 1017-1024 (1991) 1991 .John Wiley & Sons, Inc

COMPUTATIONAL DETAILS

(0

CCC 0006-3525/91/08l017-08$04.00

* Permanent address: Centro di Studio di Biocristallografia del CNH, Dipartimento di Chimica, Universitl di Napoli, via Mezzocannone 4,80134 Napoli, Italy. Permanent address: Biophysics Division, National Cancer Center Research Institute, Tsukiji, Chuo-ku, Tokyo 104,Japan.





The starting coordinates for all the simulations were those determined from the x-ray crystal s t r ~ c t u r e . ~ Energy minimizations and molecular dynamics 1017

1018

SAVIANO, AIDA, A N D CORONCIU

\

Figure 1. Comparisons of the structure of CLA in the solid state with those obtained from MD simulations: ( a ) solid state, ( b ) UNI300, ( c ) CELVAC100, ( d ) CELVAC300, ( e ) CELSOL, and ( f ) UNISOL.

( M D ) calculations were performed with the IBM 3090/400 computer using the KGNMD program from the MOTECC package,15 with the "ab initio potentials" for the solute intra- and intermolecular interactions l6 and for the water-peptide interactions, l7-I9 and with the MCY potential" for the water-water intermolecular interactions. The equations of motion were solved using the leapfrog algorithm with a time step of 0.5 fs.'l For the simulations in cell ( i n vacuo and hydrated) and in solution, the long-range Coulomb interactions were computed with the Ewald sum corrections" and bond constraints were imposed using the SHAKE algorithm.23 In all cases, periodic boundary conditions were applied.

MD Simulations In Vacuo

Two kinds of systems were simulated in vacuo: the first system was composed of one CLA molecule in a large simulation box; the second system was composed of four CLA molecules in the same crystallographic cell as in the solid state experimental data. Energy minimization using the SUMSL (secanttype unconstrained minimization solver) routine '4 was performed for one CLA molecule using the xray structure as a n initial conformation. The energyminimized structure was used as the initial structure for the MD simulations in vacuo a t 100 K (hereafter denoted a s UNIlOO) and a t 300 K (UNI300). For both simulations a n orthorombic box of 1000 X 1000

MD OF CLA

x 1000 A 3 was chosen to avoid boundary effects. For the simulations in the crystallographic cell, carried out a t 100 K (CELVAC100) and a t 300 K ( CELVAC300), the x-ray structure was used a s the starting conformation: four units per cell (2 = 4 ) were placed in a crystallographic box of 32.98 X 21.65 X 9.8 A ' with the symmetry of the space group P2,2121.In each simulation, performed with a time step of 0.5 fs, the CLA was equilibrated for 50 ps. This was followed by a n additional 60 ps of simulation without velocity rescaling, since good energy conservation was observed and the average temperature remained essentially constant around the target value of 100 K (for UNIlOO and CELVAC100) and :300 K (for UNI300 and CELVACSOO) . Coordinates and velocities for CELVAClOO and CEI,VAC300 were dumped to a disk every 10 steps during the last 20 ps of the simulations. The dumped data were used for the statistical analysis. For IJN1300, energy minimization procedures were applied several times during the equilibration phase of the MI) simulation in order to search for conformat ions closer to the global minimum. MD Simulations in Solution

Two kinds of systems were simulated: the first was composed of the CLA molecule and as many water molecules as were needed to fill the simulation box. The latter box was selected such that all peptide atoms were a t least 6 A away from the box boundaries ( CJNISOL) . The second system was composed of the same four units per cell in the crystallographic box as in the solid state data (CELSOL) and as many water molecules as were needed to fill the cell. The two simulations were performed a t a temperature of 300 K with a 0.5-fs time step. The simulations were carried out as follows: For the UNISOL case, as a first step, 640 water molecules confined in an orthorhombic box of 30 X 25 X 25 A 3 were equilibrated for a few picoseconds. As a second step, the (:LA molecule, with coordinates from the x-ray structure, was placed in the middle of the simulation box with the smallest moment of inertia parallel to the largest dimension of the box. For CELSOL, as a first step, 210 water molecules were equilibrated in an orthorhombic box of 32.98 X 21.64 X 9.83 A3. As a second step, the four units per cell were placed in the crystallographic box. Then, for both systems, we removed those water molecules with the oxygen atom within a 1.6-A distance from any peptide atom; the resulting systems were composed of 578 water molecules and 160 peptide atoms for UNISOL and of 28 water molecules and 640 peptide atoms for

1019

CELSOL. In the third step, for both systems, a 1ps simulation was performed by keeping the solute fixed a t its initial conformation to allow the water molecules to equilibrate in presence of the solute. Finally, both water and peptide atoms were allowed to move. The simulations were carried out for 30 ps in the equilibration phase and for 30 ps without velocity rescaling since the average temperature remained constant around 300 K. Coordinates and velocities for the systems were dumped to a disk every 20 steps during the last 30 ps of the two simulations. The dumped data were again used for the statistical analysis.

RESULTS MD Simulations In Vacuo

The two simulations UNIlOO and UNI300 were carried out for an isolated molecule a t 100 and 300 K, respectively, in order to obtain some insights on the characteristics of the isolated CLA molecule. In both MD simulations a t 100 and 300 K, a notable flexibility of the molecule has been observed and the rms deviations from the experimental data of the last structure of the simulations was 1.71 A a t 100 K and 2.15 A a t 300 K. T h e results are summarized in Table I, and in Figure I b we show the structure of the last conformation for the simulation a t 300 K. Table I1 compares the 6 and rC, values for the last structures of the two simulations with the solid state data. The cis bond between Pro and Pro *, present in the x-ray data, is conserved both in UNIlOO and UNI300. Large deviations, however, exist between the x-ray data and the simulation results for all dihedral angles. For UNI300 the two leucines have positive 6 and negative rC, angles, while for UNIlOO only Leu' has a negative rC, angle. Energy minimizations were repeated using as

Table I Root Mean Square Deviation (in Angstroms) from X-Ray Data for the C" Atoms of CLA, as Obtained from the MD Simulations

Single molecule (UNI100) Single molecule (UNI300) CELL vacuo (CELVAC100) CELL vacuo (CELVAC300) CELL hydrated (CELSOL) Molecule in solution (UNISOL)

T

RMS

100 300 100 300 300 300

1.71 2.15 0.35 0.83 0.62 1.41

1020

SAVIANO, AIDA, AND CORONGIU

Table I1 Comparison Between X-Ray Data and the Last Conformation of the MD Simulations for a Single Molecule at T = 100 and 300 K (UNI100 and UNI300)

X-Ray

T = 300 K

T = 100K

Residue

9

*

9

4

9

$

Pro' Pro2 Phe3 Phe4 Leu5 IIe6 lie7 Leu'

-60.0 -90.3 -97.3 -86.0 -61.5 -55.2 -114.7 55.1 -125.0

160.2 -17.7 -48.7 56.7 -28.2 -32.7 21.5 48.4 74.8

-56 -60 -124 -88 -29 -49 -120 65 -58

163 -71 34 56 -57 -31 26 -38 125

-63 -65 -75 -90 64 -158 -82 63 - 107

156 -179 61 77 -91 40 50 -53 150

va19

starting point intermediates coordinates of the MD simulation of UNI300, and many local minima were obtained with almost the same energies. T h e most stable structure is characterized by four carbonyl oxygens (Pro', Phe4, Ile7, and Val') pointing inside the ring backbone (Figure 2 ) . T h e potential energy of this conformation is 30 KJ/mol lower than the conformation of the solid state data. To study the temperature and cell-packing effect on the conformation, two other simulations, CELVAClOO and CELVAC300, were performed a t 100 and 300 K in vacuo, using the crystallographic cell as a simulation box (four units per cell). In T a bles I11 and IV the conformational statistical analysis of the two MD simulations are reported. These results have been obtained by averaging over the four units, considered a s equivalent molecules. It should be noted that the simulations have been performed without symmetry requirements within the simulation box; the space group symmetry was ap-

plied only to the periodic boundary conditions. As evident from the results in Tables I11 and IV, the two structures are similar t o the x-ray data. T h e last conformations are shown in Figure l c and Id. CELVAClOO shows the lowest rms deviation (0.35 A ) from the x-ray structure. CELVAC300 (rms = 0.83 A ) shows more flexibility around the Pro'Phe bond. MD Simulations in Solution

T h e two simulations were carried out a t 300 K: in the system UNISOL, one CLA molecule was dissolved in a box containing 578 water molecules; in the system CELSOL, four CLA molecules are packed with 28 water molecules a s in the crystalline state. In Tables V and VI, the statistical analyses obtained from the MD simulations are reported. For CELSOL the flexibility around the Pro'-Phe3 bond, already present for CELVACBOO, is increased and the rms deviation is 0.62 A. T h e last conformation is depicted in Figure le. T h e statistical analysis for the solution simulation (UNISOL) gives large differences from the x-ray structure with a n rms deviation of 1.41 A and great flexibility for all dihedral angles in the peptide main chain. In Figure lf, the last conformation is shown. In particular, the Leu' has a negative 4 and a positive $, and Leu8 a positive 4 and a negative $ with large values of deviations.

Hydrogen-Bond Analysis Figure 2. Stereo view of the most stable structure for UNI300 characterized by four carbonyl oxygens pointing inside the ring backbone. Large spheres represent these

oxygen atoms.

Hydrogen bonding plays a n important role in stabilizing the conformation of biological molecules, and in their mode of action and interaction with

MD OF CLA

1021

Table I11 Comparison of the Main Chain Dihedral Angles from X-Ray Measurements of the CLA Molecule in the Solid State with the MD Simulation in Vacuo at 100 K (CELVAC100)"

*

dJ

0

Residue

X-Ray

MD

X-Ray

MD

X - Ray

MD

Pro' Pro2 Phe:' Phe4 Leu5 Ile" Ile' Leu' Val9

-60.0 -90.3 -97.3 -98.7 -61.5 -55.2 -114.7 55.1 -125.0

-65.2 f 4.9 -71.0 f 6.7 -112.7 k 8.9 -96.7 f 7.8 -64.3 f 6.0 -43.5 f 6.7 -106.5 f 6.5 50.8 f 5.3 -127.6 f 5.7

160.2 -17.7 -48.7 56.7 -28.2 -32.7 21.5 48.4 74.8

166.1 f 4.2 -34.3 t 12.9 -16.4 ? 13.9 71.8 t 7.5 -39.3 f 5.3 -43.5 f 8.1 29.9 f 9.2 49.4 f 5.1 101.1 f 6.3

10.0 -175.9 -166.9 -179.5 178.9 179.4 175.6 169.2 175.5

-2.5 f 6.2 -174.4 f 2.5 -168.9 t 2.6 -175.3 f 2.7 177.5 f 2.8 178.4 f 2.5 179.9 t 3.4 176.5 2 2.8 167.2 f 5.5

a

We list the average of the MD results for the four units of crystallographic cell.

specific biological sites. In our analysis of the MD simulation results, we define a hydrogen bond ( H bond) when the distance between a hydrogen atom and a n acceptor atom is shorter than 2.5 A. Table VII presents the occurrence of hydrogen bonds ( % ) for the various MD simulations. We define the occurrence of hydrogen bonds as the ratio of the number of the simulation steps when the hydrogen bond is present, to the total number of simulation steps. As shown in Table VII, all hydrogen bonds found in the crystalline state are correctly reproduced in the in vacuo simulation of CLA in cells a t 100 K (CELVAC100). T h e analysis for CELVAC300 shows similar results, with the exception of the H bond between the N H group of Ile7and the CO group of P h e 4 , which is absent a t 300 K. Additional H bonds, not present in the x-ray data, have been found: between Leu5 N H and Leu' CO and between

Ile6 N H and Leu' CO, both with high probability of occurrence. By contrast, in the energy-minimized conformations of a n isolated molecule ( UNIlOO and UNI300) the hydrogen-bonding patterns are quite different from the experimental one. For these cases, we see four new H bonds: between Phe4 N H and Pro2 CO, between Val9 N H and Ile7 CO, between Leu' N H and Ile6 CO, and between Leu' NH and Phe4 CO (Figure 2 ) . In the simulation of the hydrated cell ( CELSOL) , we observe the same H-bond patterns as in the xray structure, except for the one between Phe4 N H and Val9 CO (4% occurrence is predicted by the simulation ) . Additional H bonds are observed: between Ile6 N H and Leu' CO with a high percentage of occurrence ( 7 1 % ) , and between Leu8 N H and Ile6 CO. As shown in Figure 2, the peptide-solvent hydrogen-bonding pattern in CELSOL is very close

Table IV Comparison of the Main Chain Dihedral Angles from X-Ray Measurements of the CLA Molecule in the Solid State with the MD Simulation in Vacuo at 300 K (CELVACSOO)*

*

dJ

0

Residue

X-Ray

MD

X-Ray

MD

X-Ray

MD

Pro' Pro2 Phe:' Phe4 Leu" Ile6

-60.0 -90.3 -97.3 -86.0 -61.5 -55.2 -114.7 55.1 - 125.0

-59.3 f 8.6 -70.8 f 9.1 -111.4 f 14.6 -96.7 f 11.4 -66.2 t 10.2 -42.2 f 9.7 -110.0 f 9.9 50.0 f 9.0 -127.1 f 8.3

160.2 -17.7 -48.7 56.7 -28.2 -32.7 21.5 48.4 74.8

163.3 f 7.5 -35.7 f 16.1 -15.2 k 16.6 72.9 f 10.9 -37.9 f 8.8 -48.2 k 10.7 39.4 f 8.8 51.1 t 9.1 96.4 f 8.3

10.0 -175.9 -166.9 -179.5 178.9 179.4 175.6 169.2 175.5

-3.8 f 9.7 --174.5 f 4.4 - 171.8 f 5.1 -174.1 f 4.7 179.4 f 4.6 178.1 f 4.1 177.7 2 4.5 175.6 f 4.3 165.2 f 6.7

IIe7 I,euX

ValY a

We list the average of the MD results for the four units of the crystallographic cell.

1022

SAVIANO, AIDA, AND CORONGIU

Table V Comparison of the Main Chain Dihedral Angles from X-Ray Measurements of the CLA Molecule in the Solid State with the MD Simulation in Solution at 300 K (CELSOL)"

G

4

w

Residue

X-Ray

MD

X-Ray

MD

X-Ray

MD

Pro' Pro2 Phe3 Phe4 Leu' Ile6 1ie7 Leu8 va19

-60.0 -90.3 -97.3 -86.0 -61.5 -55.2 -114.7 55.1 -125.0

-58.8 f 8.1 -69.1 i 11.8 -117.1 f 12.4 -75.7 f 7.4 -59.8 f 9.7 -42.5 f 11.3 -102.8 f 11.2 48.5 f 10.4 -124.2 i 9.0

160.2 -17.7 -48.7 56.7 -28.2 -32.7 21.5 48.4 74.8

161.5 i 7.8 -42.8 f 17.2 -23.9 i 12.5 63.4 f 9.3 -34.9 f 10.1 -44.1 f 11.2 32.1 f 12.2 47.5 f 9.4 100.9 i 8.1

10.0 -175.9 -166.9 -179.5 178.9 179.4 175.6 169.2 175.5

-3.6 k 9.1 175.6 f 5.0 179.5 i 5.0 -177.1 f 4.6 176.4 f 4.5 178.1 f 4.2 -178.3 f 4.8 176.8 f 4.8 161.6 f 6.6

a

We list the average of the MD results for the four units of crystallographic cell.

t o that found in the solid state7 with three extra peptide-solvent H bonds: between the oxygen of one water molecule and the CO of Pro', between Phe4 N H and 0 ( w ) , and between Leu' N H and 0 ( w ) . In the solution simulation (UNISOL) , only three of the H bonds found in the solid state are observed between Leu5 N H and Phe3 CO, between Ile7 N H and Phe4 CO, and between Leua N H and Leu5 CO, all with a low percentage of occurrence. However, there are new hydrogen bonds: between Val9 NH and Ile7 NH, with a high percentage of occurrence (93%), and between Ile6 N H and Phe4 CO.

DISCUSSION We have performed MD simulations of the CLA molecule in various environments. T h e analysis of the simulations shows the possibility of existence of various structures depending upon the molecular

environment. The results are comparable with experimental observations. T h e MD simulation of UNISOL was performed in order t o analyze the conformational characteristics of the CLA molecule in a polar solvent: in such a n environment it has been impossible to resolve the structure using nmr techniques. The simulation result a t 300 K points out that CLA is not well structured in water, namely, we do not find any single well-defined conformation. This is in good agreement with experimental observations, where one observes the existence, in solution a t room temperature, of several conformers. T h e MD simulations of CELVAClOO and CELVAC300 were performed in order to study the packing effect. I t is noteworthy that the MD results of CELVAClOO ( t h e system composed only of four CLA molecules ) show structural characteristics similar to those of the x-ray and nmr data in chloroform a t 214 K. whereas those for CLA in solution

Table VI Averages and Relative Errors of the Dihedral Angles in the Peptide Main Chain as Obtained from the MD Simulation in Solution at 300 K (UNISOL) Residue

4

G

w

Pro' Pro2 Phe3 Phe4 Leu6 Ile6

-59.1 f 7.5 -87.3 2 8.1 -104.3 k 11.3 -113.1 2 14.8 -68.3 f 14.5 -104.4 2 40.8 -126.8 f 17.0 61.5 f 9.0 -65.2 f 21.7

177.6 2 6.4 -40.3 f 17.5 -9.3 f 19.7 94.1 f 13.5 14.9 -t 48.4 -17.1 f 29.6 55.2 f 15.5 -49.2 f 24.5 124.6 i 8.9

-14.1 f 10.3 -172.9 f 4.6 178.8 -t 5.6 -176.5 f 4.7 -178.0 i 5.5 -177.5 i 4.9 179.9 f 4.6 -177.6 f 4.8 152.9 -t 7.8

1ie7

Leu' va19

1023

MD OF CLA

Table VII The Occurrence of Intramolecular Hydrogen Bonds ( X ) Obtained from MD Simulations Compared with Those from X-Ray Data7 CELL Vacuum

T

NH-D

CO-A

X-ray

va19 Phe4 Leus 1ie7 Leu' I , ~ Ile' LeuR va19 Ile6 1ie7 va19

Phe4 va19 Phe3 Phe4 Leu5 ~Leu'~ Leu8 IIe6 Leu' Phe4 Leu' 1ie7

100 100 100 100 100

100 99.5 70 20 100

-

-

=

100K

(UNISOL) are quite different from the x-ray data. This indicates that the packing of the CLA molecules in the cell plays a n important role in determining the peptide conformation of the crystallographic environment. T h e MD simulation of CELSOL was performed in order to analyze the packing effect in presence of water molecules, a t 300 K. The analyses show the

T

=

CELL Hydrated Solution 300 K

97 92 66 1 95 36 39 3 -

T

=

300 K

60 4 99.7 32 99.6

T

=

300 K

-

4 19 5

4

-

71 20 2 -

-

4 1 30 4 93

existence of the same solvent-peptide hydrogen bonds as in the experimental data; however, additional hydrogen bonds between CLA and water are detected. T h e solid state conformational study was performed with a crystal grown from 2-propanolwater mixtures. The flexibility of the Xxx-Pro bonds, observed in the nmr study, can be explained by the tendency of a hydrogen bond to form between

,

.

Figure 3. Arrangement of the CLA and water molecules after MD simulation at 300 K (CELSOL). Only the hydrogen bonds between peptide and solvent, already found in the x-ray structure, are shown as dashed lines.

1024

SAVIANO, AIDA, A N D CORONGIU

the oxygen of one water molecule and Pro :the N H and CO groups can be involved in both intramolecular and solvent-peptide hydrogen bonds. The presence of the peptide-water H bond between the N H of P h e 4 and the oxygen of one water molecule can be a possible explanation for the low percentage of occurrence of the H bond between Phe4 N H and Val9 CO (Table VII). It should be emphasized that the arrangement of the water molecules found in CELSOL (Figure 3 ) is quite similar to the one found in the solvent molecules in the solid state study.’ T h e water molecules fill a channel in the c crystallographic axis direction, bridging molecules of CLA located in a layer perpendicular to the b axis. These layers of CLA and solvent molecules are held together by van der Waals contacts between interacting side-chain hydrophobic groups. The MD simulations of UNIlOO and UNI300 and further energy minimizations were performed in order to clarify the characteristics of one CLA molecule in vacuo. T h e results point out the existence of different conformations with almost the same energies. T h e flexibility of this molecule supports the possibility of complex formation between CLA and metal ions in solution, as already pointed out in Ref. 25.

CONCLUSIONS In the present study we have investigated the characteristics of the CLA molecule both in vacuo and in solution, and have compared our results with those observed experimentally in solution and in the solid state. T h e MD results show that this cyclic molecule in water exists in several conformations with high flexibility of the two Xxx-Pro bonds, and with the capability of the solvent water molecules to divert the NH and CO groups from the formation of intramolecular hydrogen bonds. T h e results of the MD simulations of CLA in the cell environment are in agreement with the x-ray data and nmr data in chloroform a t low temperature. This result emphasizes the role of the packing effect in determining the peptide conformation in the x-ray experiments. We thank Profs. C. Pedone and E. Benedetti for their suggestions and for proposing this research.

REFERENCES

J. W., Ziegler, K. & Frimmer, M. (1986) Angew. Chem. Int. 25,997-999. 3. Wieland, T . ( 1972) in Chemistry and Biology of Peptides, Meinhofer, J., Ed., Ann Arbor Science, Ann Arbor, MI, pp. 377-396. 4. Kessler, H., Bats, J. W., Griesinger, C., Koll, S., Will, M. & Wagner, K. J. (1986) A m . Chem. SOC.110, 1033-1049. 5. Di Blasio, B., Benedetti, E., Pavone, V., Pedone, C. & Goodman, M. (1987) Biopolymers 26,2099-2101. 6. Tancredi, T., Zanotti, G., Rossi, F., Benedetti, E., Pedone, C. & Temussi, P. A. (1989) Biopolymers 28, 513-523. 7. Di Blasio, B., Rossi, F., Benedetti, E., Pavone, V., Pedone, C., Temussi, P. A., Zanotti, G. & Tancredi, T. (1989) J. A m . Chem. SOC.111,9089-9098. 8. Naider, F., Benedetti, E. & Goodman, M. (1971) Proc. Natl. Acad. Sci. 68, 1195-1198. 9. Brewster, A. I. & Bovey, F. A. (1971) Proc. Natl. Acad. Sci. 68, 1199-1202. 10. Siemion, I. Z., Klis, W. A., Sucharda-Sobczyk, A. & Obermeier, R. (1977) Rocniki Chem. 51,1489-1498. 11. Tonelli, A. E. (1971) Proc. Natl. Acad. Sci. 68, 12031207. 12. Karle, I. L., Karle, J., Wieland, T., Burgermeister, W., Faulstich, H. & Witkop, B. (1973) Proc. Natl. Acad. Sci. 6, 1836-1840. 13. Karle, I. L., Wieland, T., Schermer, D. & Ottenheym, H. C. (1979) Proc. Natl. Acad. Sci. 76,1532-1536. 14. Castiglione Morelli, M. A., Pastore, A., Pedone, C., Temussi, P. A., Zanotti, G. & Tancredi, T. ( 1990) Int. J . Peptide Protein Res., in press. 15. Clementi, E., Corongiu, G., Aida, M., Niesar, U. & Kneller, G. (1990) in Modern Techniques i n Computational Chemistry: MOTECC-90, Clementi, E., Ed., ESCOM Publishers, Leiden, chap. 17. 16. Aida, M., Corongiu, G. & Clementi, E., to be published. 17. Clementi, E., Cavallone, F. & Scordamaglia, E. (1977) J. Am. Chem. SOC.99, 5531. 18. Ranghino, G., Clementi, E. & Romano, S. (1983) Biopolymers 22, 1449. 19. Ragazzi, M., Ferro, S. & Clementi, E. (1979) J . Chem. Phys. 70,1040. 20. Matsuoka, O., Clementi, E. & Yoshimine, M. (1976) J . Chem. Phys. 64, 1351. 21. Hockney, R. W. (1970) Methods Comput. Phys. 9, 136. 22. De Leeuw, S. W., Perran, J. W. & Smith, E. R. (1980) Proc. Roy. Soc. (London) A 373,27. 23. Rychaert, J. P., Ciccotti, G. & Berendsen, H. J. C. (1977) J. Comput. Phys. 23,327. 24. Dennis, J. E., Gay, D. M. & Welsch, R. E. (1981) AC M Trans. Math. Software 7,369. 25. Rossi, F., Saviano, M., Benedetti, E., Pedone, C., Temussi, P. A,, Zanotti, G. & Tancredi, T. (1990) Biopolymers, in press.

1. Kaufmann, H. P. & Tobschirbel, A. (1959) Chem. Ber.

92,2805-2809. 2. Kessler, H., Klein, M., Muller, A., Wagner, K., Bats,

Received February 7, 1991 Accepted April 29, I991

Molecular dynamics simulation in vacuo and in solution of cyclolinopeptide A: a conformational study.

The conformation of cyclolinopeptide A [c-(Pro-Pro-Phe-Phe-Leu-Ile-Ile-Leu-Val)], a naturally occurring peptide with remarkable cytoprotective activit...
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