Crystals of the title compound, C20,H29,N3,O6, are monoclinic, space group P2, with a = 8-839 (3), b = f10.818 (3), c = 11.414 (2) A, &#946; = 95.69 (2)&#176; Z = 2; final R = 0.053. The molecular conformation is defined by the following angles (&#966;, &#968;): Aib-1 58- 1, 36.8; Aib-2 68.3, 18.6; Ala-3 (&#966;) -136.2&#176;. The molecule adopts a type 111 &#946; -turn conformation stabilized by an intramolecular hydrogen bond between the CO of the benzyloxycarbonyl group and the NH of the alanyl residue. The hydrogen-bond parameters are N&#183;&#183;&#183;O 2-904 &#197; and &#8736;NH&#183;&#183;&#183;O 156.9&#176;

Balaram, P.

Nagaraj, R.

Shamala, N.

Venkataram Prasad, B. V.

English

Publications of the IAS Fellows

The Crystal Structure of Benzyloxycarbonyl-( a-aminoisobutyryl),-L-Alanyl Methyl Ester 
BY B. V. VENKATARAM PRASAD, N. SHAMALA, R. NAGARAJ AND P. BALARAM 
Molecular Biophysics Unit, Indian Institute of Science, Bangalore-560 0 12, India 
Abstract 
Crystals of the title compound, C,,H,,N,O,, are 
monoclinic, space group P2, ,  with a = 8-839 (3), b = 
final R = 0.053. The molecular conformation is defined 
by the following angles (q, y): Aib-1 58- 1, 36.8; Aib-2 
68.3, 18.6; Ala-3 (q) -136.2O. The molecule adopts a 
type 111’ p-turn conformation stabilized by an intra- 
molecular hydrogen bond between the CO of the 
benzyloxycarbonyl group and the NH of the alanyl 
residue. The hydrogen-bond parameters are N .O 
2-904 A and LNH. . .O  156.9O. 
10.818 (3), c = 11.414 (2) A, p = 95.69 (2)O, 2 = 2; 
Introduction 
a-Aminoisobutyric acid (Aib) is an important con- 
stituent of the membrane-modifying polypeptide 
alamethicin and related microbial peptides (Martin & 
Williams, 1976; Pandey, Carter Cook & Rinehart, 
1977). The conformational behaviour of Aib- 
containing peptides has been studied to establish 
conformation-function correlations for these poly- 
peptides. The presence of gem-dialkyl substituents at 
the Ca atom greatly restricts conformational freedom 
at Aib residues and thus permits the observation of well 
defined conformations in solution, which may then be 
compared with the solid-state structures (Nagaraj, 
Shamala & Balaram, 1979). In this paper we describe 
the molecular structure of the tripeptide ester 
benzyloxycarbonyl-Aib-Aib-Ala-OMe, which is shown 
to adopt a type 111’ p-turn structure, stabilized by a 
4+ 1 intramolecular hydrogen bond. 
Experimental 
2-Aib-Aib-Ala-OMe was synthesized from 2-Aib-Aib- 
OH and Ala-OMe with dicyclohexylcarbodiimide, 
followed by work up (Nagaraj, Shamala & Balaram, 
1979). 
Crystals of 2-Aib-Aib-Ala-OMe (C,,H,,N,O,, Mr = 
407) belonging to the monoclinic system were obtained 
by slow evaporation of a chloroform solution. Cell 
dimensions were obtained from 20  measurements both 
on photographs and on a diffractometer. Crystal data 
are: space group P2,,  a = 8.839 (3), b = 10.818 (3), 
The density determined in a KC1-water mixture by 
flotation is 1.23 Mg m-3 and the calculated density is 
1.24 Mg m-3. Intensities of 2655 reflections were 
measured on a CAD-4 diffractometer with Cu Ka 
radiation and an w 2 8  scan. Two standard reflections 
were measured after every 100 reflections and there 
was no significant change in their intensities. The 
intensities were corrected for Lorentz and polarization 
factors but not for absorption, since the crystal was 0.4 
x 0.3 x 0.1 mm. 
c = 11.414 (2) A, p = 95.69 (2)O, Z = 2, V =  1086 A3. 
Structure determination and refinement 
The structure was solved by direct methods (Karle & 
Karle, 1966) with MULTAN (Germain, Main & 
Woolfson, 197 1). The normalized structure factors 
(Karle & Hauptman, 1956) were obtained with the 
overall temperature factor (B = 3.36 A2) and scale 
factor (0.129) determined from a Wilson (1 942) plot. E 
maps were computed with 300 reflections having E 2 
1.5. Twenty-three non-hydrogen atoms out of 29 could 
be located in an E map corresponding to the highest 
figure of merit. Calculated structure factors for the trial 
coordinates of the partial structure gave R = 0.41. 
After refinement of positional and isotropic thermal 
parameters by block-diagonal least squares (R. Shiono, 
personal communication) R converged to 0.36. A 
difference map computed at  this stage revealed the 
positions of the remaining six atoms. Further refine- 
ment lowered R to 0.146. Scattering factors for non- 
hydrogen atoms were from Cromer & Waber (1965). 
Table 2. Positional ( x  lo3) and isotropic thermal ( x  10) 
parameters of the hydrogen atoms 
Refinement of all non-hydrogen atoms with aniso- 
tropic temperature factors yielded R = 0.096. The 
positions of the 20 H atoms could be obtained from a 
difference map but all the H atoms attached to C atoms 
were fixed with C-H = 1.1 A. Bond angles of 109.5O 
or 120-0° were used for tetrahedral and trigonal atoms, 
respectively. For H bonded to N atoms, N-H = 1 .O 8, 
and a bond angle of 120-0° were used. The H atoms 
were assigned the temperature factor of the carrier 
atom. With the scattering factor for H given by 
Stewart, Davidson & Simpson (1965), the refinement 
of positional and anisotropic thermal parameters of 
non-hydrogen atoms and positional and isotropic 
thermal parameters of H atoms yielded an R of 0.069. 
With the weighting scheme of Cruickshank (1961) 
further refinement converged to a final R of 0.053 for 
2643 reflections. The final difference map was feature- 
less. The shifts in the parameters at the end of the last 
cycle were (0- la. The atomic parameters are given in 
Tables 1 and 2.* 
* Lists of structure factors and anisotropic thermal parameters 
have been deposited with the British Library Lending Division as 
Supplementary Publication No. SUP 34782 (16 pp.). Copies may 
be obtained through The Executive Secretary, International Union 
of Crystallography, 5 Abbey Square, Chester CH 1 2HU, England. 
Table 1. Positional coordinates for the non-hydrogen 
atoms (x  lo4) 
E.s.d.’s are given in parentheses. 
X 
2847 (3) 
3677 (3) 
4812 (4) 
5094 (4) 
4286 (3) 
3 140 (3) 
2164 (3) 
3062 (2) 
3202 (2) 
2605 (2) 
4042 (2) 
4623 (2) 
5792 (3) 
5367 (3) 
3327 (2) 
3543 (2) 
2021 (2) 
715 (2) 
1183 (3) 
-477 (3) 
-17 (2) 
-938 (2) 
293 (2) 
-273 (3) 
-845 (3) 
-415 (3) 
-1880 (3) 
-2540 (6) 
933 (4) 
Y 
4976 (3) 
5085 (3) 
4235 (4) 
3281 (4) 
3 180 (0) 
3995 (3) 
3816 (3) 
3961 (2) 
5138 (2) 
6018 (2) 
5 185 (2) 
6361 (2) 
6908 (3) 
6089 (2) 
7282 (2) 
8395 (2) 
6827 (2) 
76 10 (2) 
8583 (3) 
6762 (3) 
8231 (2) 
9087 (2) 
7735 (2) 
8250 (3) 
9008 (4) 
7 184 (3) 
7012 (4) 
65 19 (3) 
5507 (5) 
Z 
-78 (2) 
Bonded 
to 
H(1) C(1) 
H(2) C(2) 
H(3) (33) 
H(4) C(4) 
H(5) C(5) 
H(6) C(7) 
H(7) C(7) 
H(8) N(1) 
H(9) C(10) 
H(10) C(l0) 
H( l  1)  C(10) 
H(12) C(11) 
H(13) C(11) 
H(14) C(11) 
H(15) N(2) 
H(16) C(14) 
H(17) C(14) 
H(18) C(14) 
H(19) C(15) 
H(20) C(15) 
H(21) C(15) 
H(22) N(3) 
H(23) C(17) 
H(24) C(18) 
H(25) C(18) 
H(26) C(18) 
H(27) C(20) 
H(28) C(20) 
H(29) C(20) 
E.s.d.’s are given in parentheses. 
X 
207 ( 5 )  
331 (7) 
551 (4) 
600 ( 5 )  
457 (6) 
121 (4) 
173 (5) 
431 (4) 
517 ( 5 )  
628 (4) 
663 (4) 
619 (4) 
583 (5) 
453 (3) 
216 (6) 
204 (4) 
2 ( 5 )  
183 (8) 
-22 (4) 
- 140 (5) 
-73 ( 5 )  
97 (4) 
-64 (9) 
197 (5) 
62 ( 5 )  
121 (6) 
-175 (7) 
-328 (6) 
-289 (6) 
Y z 
556 ( 5 )  6 (4) 
565 (7) -170 (5) 
428 (4) -184 (3) 
275 (6) -72 (4) 
244 (6) 118 ( 5 )  
440 (5) 165 (3) 
286 (6) 180 (4) 
456 (4) 446 (3) 
706 (6) 300 (4) 
778 ( 5 )  429 (3) 
635 (4) 399 (3) 
555 (5) 608 (3) 
685 (6) 639 (4) 
569 (4) 655 (3) 
626 (6) 553 (4) 
900 (4) 632 (3) 
913 (5) 657 (4) 
824 (9) 721 (6) 
641 (4) 672 (3) 
713 (6) 640 (4) 
627 ( 5 )  550 (3) 
717 (4) 337 (3) 
904 (10) 844 (6) 
837 (6) 151 (4) 
953 ( 5 ;  95 (4) 
965 (7) 229 (5) 
481 (8) 94 (5) 
516 (7) 158 ( 5 )  
592 (6) 36 (4) 
Results and discussion 
B (A2) 
57 (9) 
86 (15) 
39 (8) 
62 (13) 
70 (12) 
47 (9) 
68 (10) 
27 (7) 
66 (11) 
50 (9) 
36 (7) 
43 (9) 
44 (10) 
30 (6) 
40 (8) 
57 (10) 
125 (18) 
38 (7) 
60 (10) 
39 (8) 
35 (8) 
112 (24) 
53 (10) 
49 (9) 
86 (14) 
92 (15) 
90 (13) 
77 (12) 
68 (12) 
-1050 (2) 
-1234 (2) 
-466 (3) 
513 (2) 
A perspective view of the molecule down a is shown in 
Fig. 1. Bond lengths and angles are listed in Table 3. 
The bond lengths and angles of the peptide units 
706 (2) 
1709 (2) 
2847 (1) 
3236 (2) 
2719 (1) 
4289 (1) 
4786 (2) 
4020 (2) 
6039 (2) 
4928 (2) 
4785 (2) 
5251 (1) 
5499 (2) 
6445 (2) 
5957 (2) 
4365 (2) 
4433 (2) 
3347 (2) 
2209 (2) 
1678 (3) 
1393 (2) 
445 (2) 
1853 (2) 
1147 (4) 
Fig. 1. Molecular conformation of Z-Aib-Aib-Ala-OMe 
down a. 
viewed 
Table 3. Bond distances (A) and angles (O) with e.s.d.S 
in parentheses 
C(l)-C(2) 1.395 (4) 
C(2)-C(3) 1.392 ( 5 )  
C(3)-C(4) 1.361 ( 5 )  
C(4)-C(5) 1.389 (4) 
C(5)-C(6) 1.377 (4) 
C(6)-C( 1) 1.396 (4) 
C(6)-C(7) 1.5 13 (3) 
C(7)-O( 1) 1.462 (3) 
O( 1)-C(8) 1.350 (3) 
C(8)-0(2) 1.213 (3) 
C(S)-N( 1) 1.350 (3) 
N( 1)-C(9) 1.465 (3) 
C(9)-C( 10) 1.536 (3) 
C(9)-C( 11) 1.543 (3) 
C(9)-C(12) 1.540 (3) 
C( l)-C(2)-C(3) 120.6 (3) 
C(2)-C(3)-C(4) 119.5 (3) 
C(4)-C(5)-C(6) 121.6 (3) 
C(5)-C(6)-C( 1) 118.5 (2) 
C(6)-C( 1)-C(2) 119.7 (3) 
C(5)-C(6)-C(7) 121.5 (2) 
C( l)-C(6)-C(7) 119.9 (2) 
C(6)-C(7)-0( 1) 110.9 (2) 
C(7)-O( 1)-C(8) 114.7 (2) 
O( 1)-C(8)-0(2) 124.0 (2) 
O( l)-C(S)-N( 1) 110.7 (2) 
C(8)-N( 1)-C(9) 12 1.2 (2) 
0(2)-C(S)-N( 1) 125.3 (2) 
N( l)-C(9)-C( 10) 110.2 (2) 
N( 1)-C(9)-C( 11) 107.1 (2) 
N(l)--C(9)-C(12) 1 1  1.5 (2) 
C( lO)-C(9)-C( 11) 110.6 (2) 
C( lo)-C(9)-c( 12) 11 1 * 1 (2) 
C(3)-C(4)-C(5) 120.1 (3) 
C( 11)-C(9)-C( 12) 106.2 (2) 
C(9)-C(12)-N(2) 117.5 (2) 
C(9)-C( 12)-O(3) 119.4 (2) 
C( 12)-N(2) 
C( 12)-0(3) 
N(2)-C( 13) 
C( 13)-C( 14) 
C( 13)-C( 15) 
C( 13)-C( 16) 
C ( 16)-0(4) 
N(3)-C( 17) 
C( 17)-C( 18) 
C( 17)-C( 19) 
C( 19)-O(5) 
c ( 19>-0(6) 
0(6)-C(20) 
C(16)-N(3) 
1.339 (3) 
1.233 (3) 
1.481 (3) 
1.535 (4) 
1.528 (4) 
1.543 (3) 
1.240 (3) 
1.333 (3) 
1.456 (3) 
1.518 ( 5 )  
1.536 (4) 
1.196 (4) 
1.313 (4) 
1.448 (6) 
O(3)-C( 12)-N(2) 
C( 12)-N(2)-C( 13) 
0(5)-C( 19)-0(6) 
C( 17)-C( 19)-0(6) 
N(2)-C( 13)-C( 14) 
N(2)-C(13)-C(15) 
N(2)-C( 13)-C( 16) 
C(15)-C(13)-C(14) 
C( 15)-C( 13)-C ( 16) 
C( 14)-C( 13)-C ( 16) 
C ( 13)-C ( 16)-N (3) 
C( 13)-C( 16)-0(4) 
O(4)-C( 16)-N(3) 
C( 16)-N(3)-C( 17) 
N(3)-C( 17)-C( 19) 
N(3)-C(17)-C( 18) 
C(18)-C(17)-C(19) 
C( 17)-C( 19)-O(5) 
C( 19)-0(6)-C(20) 
123.0 (2) 
123.5 (2) 
124.9 (3) 
1 1  1.7 (3) 
1 1  1.4 (2) 
107.4 (2) 
11  1 *o (2) 
108.6 (2) 
107.6 (2) 
110.6 (2) 
116.7 (2) 
119.7 (2) 
123.3 (2) 
122.8 (2) 
108.5 (2) 
11 1.8 (3) 
1 I 1.4 (3) 
123.4 (3) 
116.7 (3) 
compare well with those found in other peptides 
(Marsh & Donohue, 1967; Ramachandran, Kolaskar, 
Ramakrishnan & Sasisekharan, 1974; Benedetti, 
1977). The average C-H = 1.0 A and N-H = 0-8 A. 
The angles involving H with tetrahedral C atoms are 
around 109O on average. There are few intramolecular 
contacts less than the normal limits given by 
Ramachandran & Sasisekharan (1968). The contacts 
C(7)-0(2) of 2.659(4) A and C(20)-0(5) of 
2.668 (6) 8, are less than the proposed extreme limit of 
The molecular conformation is stabilized by an 
internal hydrogen bond between the CO of the urethan 
moiety and the N H  of the alanyl residue. This 
conformation with a 4-1 type of intramolecular 
hydrogen bond, which corresponds to a type 111’ p- 
bend (Venkatachalam, 1968), has also been seen earlier 
in oligopeptides containing Aib residues (Shamala, 
Nagaraj & Balaram, 1977, 1978; Prasad, Shamala, 
Nagaraj, C handrasekaran & Balaram, 1979; Smith, 
2 .7  A. 
Duax, Czerwinski, Kendrick, Marshall & Mathews, 
1977). The conformational angles of the backbone are 
listed in Table 4. All the peptide units are trans and 
only small deviations of about 5 O  from planarity are 
observed. 
Theoretical calculations suggest that the additional 
methyl substituent at Ca of the Aib residue greatly 
restricts the allowed values of (Q and y to the right- and 
left-handed 3 ,o- and a-helical regions (Marshall & 
Bosshard, 1972; Burgess & Leach, 1973). The values 
in Table 5 show that both the Aib residues occur in the 
left-handed helical region of the conformational map. 
In peptides containing L amino acids like Z-Aib-Pro- 
Aib- Ala-OMe (Nagaraj et al., 1979), Boc-Pro-Aib-Ala- 
Aib-OMe (Smith et al., 1977) and Z-Aib-Pro-NHMe 
(Prasad et al., 1979) the q, y values for the Aib 
residues fall in the right-handed helical region. How- 
ever, in tosyl-(Aib),-OMe (Shamala et al., 1977) both 
helical senses are equally probable as observed in the 
crystal, which belongs to a centrosymmetric space 
group. It is clear that the presence of an Aib residue 
Table 4. Conformational angles (O) for  the peptide 
backbone according to IUPA C-IUB Commission on 
Biochemical Nomenclature ( 1970) 
w,[O( 1)-C (8)-N ( 1 )-C (9)I 
p2[ C (8)-N( 1)-C (9)-C ( 12)1 
I,U?[ N( I )-C (9)-C ( 12)-N (211 
~,[C(9)-C(l2)-N(2)-C(13)1 
p3[ C ( 12)-N(2)-C( 13)-C ( 16)I 
I+Y.,[ N( 2)-C( 13)-C( 16)-N(4)I 
W,[C( 13)-C( 16)-N(4)-C( 17)I 
p4[ C ( 16)-N (4)-C( 1 7)-C ( 1 8)I 
167.2 (2) 
58.1 (2) 
36.8 (2) 
175.8 (2) 
68.3 (2) 
18.6 (3) 
-177.7 (2) 
-136.2 (2) 
J 
Fig. 2. Packing diagram viewed down b (--- intermolecular 
hydrogen bonds; -.-.- intramolecular hydrogen bonds). 
Table 5.  Details of the hydrogen bonds 
Donor Acceptor 
D A D * . . A  L D - H * * . A  L H - D . * . A  
N(3) 0(2)* 2.904 (7) A 156-9 (25)' 16.5 (20)' 
N(1) O(3Y 2.997 (7) 144.5 (24) 27.2 (21) 
N(2) O(4)" 3.147 (8) 138.7 (24) 32.9 (22) 
Symmetry code: superscript: (i) 1 - x, 1 - (y + 9, 1 - t; (ii) 
--x, 1 - (y + f), 1 - 2. 
* Intramolecular hydrogen bond. 
induces fibend formation and the consequent 
generation of a 3 ,,-helical segment. 
The crystal structure viewed along b is shown in Fig. 
2. In addition to the good intramolecular NH.m-0 
hydrogen bond between the CO of the urethan moiety 
and NH of the alanyl residue, the other peptide NH and 
C 0 groups are involved in intermolecular hydrogen 
bonds. The details of the inter- and intramolecular 
hydrogen bonds are given in Table 5 .  
This research was supported by the University 
Grants Commission. NS thanks the CSIR for financial 
support. RN was the recipient of a fellowship from the 
Department of Atomic Energy. 
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The crystal structure of benzyloxycarbonyl-(α-aminoisobutyryl)-L-alanyl methyl ester

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The crystal structure of benzyloxycarbonyl-(α-aminoisobutyryl)-L-alanyl methyl ester

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