Nucleic Acids Research, Vol. 18, No. 15 4601
. 1990 Oxford University Press
Nucleotide
of
sequence
cDNA encoding ammodytin L
a
Joze Pungercar, Ningsheng Liang*, Borut Strukelj and Franc Gubensek Department of Biochemistry, Jozef Stefan Institute, Jamova 39, 61 11 1 Ljubljana, Yugoslavia Submitted June 27, 1990
EMBL accession no. X53036
ACKNOWLEDGEMENTS
Ammodytin L is a very basic phospholipase A2 homologue isolated from European viper (Vipera ammodytes) venom as the fraction 1 (1). In contrast to other highly presynaptically toxic phospholipases A2 from the same animal ammodytoxins A, B and C (2-4), it is presumably non-neurotoxic and lacks enzyme activity. However, a significant myotoxic activity has been reported recently (5). A cDNA coding for ammodytin L was found by screening a venom gland cDNA library with a mixed oligonucleotide probe deduced from the part of a highly conserved region responsible for calcium- binding found in most phospholipases A2. It codes for a 122 amino acid residue mature protein, preceded by a 16 residue signal peptide (underlined). Ammodytin L cDNA shows about 90% nucleotide and 74% amino acid similarity to ammodytoxins C and B (6, 7). One of the most interesting amino acid replacements is the substitution of aspartic acid for the serine at position 48. Asp-48 is essential for the binding of calcium ion and therefore for enzymatic activity (8). Additionally, microheterogeneity was found by protein sequencing of ammodytin L at position 71 where proline and serine were detected (9).
This work was supported by the grants of the Research Community of Slovenia and Federal Secretariat for Development.
REFERENCES 1. Guben§ek,F., Ritonja,A., Zupan,J. and Turk,V. (1980) Period. Biol. 82, 443-447. 2. Ritonja,A. and Gubengek,F. (1985) Biochim. Biophys. Acta 828, 306-312. 3. Ritonja,A., Machleidt,W., Turk,V. and Guben§ek,F. (1986) Bio. Chem.
Hoppe-Seyler 367, 919-923. 4. Kriiaj,I., Ritonja,A., Turk,D. and Gubensek,F. (1989) Biochem. Biophys. Acta 999, 198-202. 5. Bieber,A.L. personal communication. 6. Punger6ar,J., Kordis,D., Jerala,R., Trstenjak- Prebanda,M., Dolinar,M., Curin-Serbec,V., Komel,R. and Gubensek,F. (1989) Nucl. Acids Res. 17, 4367. 7. Kordis,D., Pungercar,J., 9tnukelj,B., Liang,N. and Gubensek,F. (1990) Nucl. Acids Res. 18, 4016. 8. van den Berg,C.J., Slotboom,A.J., Verheij,H.M. and de Haas,G.H. (1989) J. Cell. Biochem. 39, 379-390. 9. Kriiaj,I., Ritonja,A. and Gubensek,F. (1990) in preparation.
TGGACCCCCTTCAACTCTGAGACAAGGCTGCCAGCTGTCTGGATTCAGGAGG
-1
ATGAGGATTCTCTGGATAGTGGCCGTGTGCCTGATAGGCGTCGAGGGGAGCGTGATCGAA
60
M
R
I
L
W
I
V
A
V
C
L
I
G
V
E
G
S
V
I
20
E
TTTGGGAAGATGATCCAGGAGGAGACGGACAAAAACCCTCTTACCTCCTACTCCTTTTAC F
G
K
M
I
Q
E
E
T
D
K
N
P
L
T
S
Y
S
F
Y
120 40
G
C
H
C
G
L
G
N
K
G
K
P
K
D
A
T
D
R
C
C
180 60
TTCGTGCATAGCTGCTGTTACGCGAAACTGTCCGACTGCAGTCCCAAAACGAACAGATAC F
V
H
S
C
C
Y
A
K
L
S
D
C
S
P
K
T
N
R
GAATACCACCGGGAGAACGGGGCTATCGTCTGTGGAAGCAGCACCCCGTGCAAGAAACAG E
Y
H
R
E
N
G
A
I
V
C
G
S
S
T
P
C
K
K
240
80
Y
Q
300 100
360 I
C
E
C
D
R
A
A
A
I
C
F
R
E
N
L
K
T
Y
N
AAGAAATATAAGGTGTACCTGAGGTTTAAATGCAAGGGGGTTTCAGAGAAATGCTAAGTC K
K
Y
K
V
Y
L
R
F
K
C
K
G
V
S
E
K
C
*
120
420
138
TCTGCAGGCCGGGGAAAAACCCTCCAATTACACAATCGTAGTTGTGTTACTCTATTATTC 480 TGAATGCAATACTGAGCAATAAACAGGTGTCAGCTCTACACTCAAAAAAAAAAAAAAAAAA 541
*Permanent address: Laboratory of Pharmacology, Cancer Institute, Medical College, PO Box 530027, Nanning, China
. 1990 Oxford University Press
Nucleotide
of
sequence
cDNA encoding ammodytin L
a
Joze Pungercar, Ningsheng Liang*, Borut Strukelj and Franc Gubensek Department of Biochemistry, Jozef Stefan Institute, Jamova 39, 61 11 1 Ljubljana, Yugoslavia Submitted June 27, 1990
EMBL accession no. X53036
ACKNOWLEDGEMENTS
Ammodytin L is a very basic phospholipase A2 homologue isolated from European viper (Vipera ammodytes) venom as the fraction 1 (1). In contrast to other highly presynaptically toxic phospholipases A2 from the same animal ammodytoxins A, B and C (2-4), it is presumably non-neurotoxic and lacks enzyme activity. However, a significant myotoxic activity has been reported recently (5). A cDNA coding for ammodytin L was found by screening a venom gland cDNA library with a mixed oligonucleotide probe deduced from the part of a highly conserved region responsible for calcium- binding found in most phospholipases A2. It codes for a 122 amino acid residue mature protein, preceded by a 16 residue signal peptide (underlined). Ammodytin L cDNA shows about 90% nucleotide and 74% amino acid similarity to ammodytoxins C and B (6, 7). One of the most interesting amino acid replacements is the substitution of aspartic acid for the serine at position 48. Asp-48 is essential for the binding of calcium ion and therefore for enzymatic activity (8). Additionally, microheterogeneity was found by protein sequencing of ammodytin L at position 71 where proline and serine were detected (9).
This work was supported by the grants of the Research Community of Slovenia and Federal Secretariat for Development.
REFERENCES 1. Guben§ek,F., Ritonja,A., Zupan,J. and Turk,V. (1980) Period. Biol. 82, 443-447. 2. Ritonja,A. and Gubengek,F. (1985) Biochim. Biophys. Acta 828, 306-312. 3. Ritonja,A., Machleidt,W., Turk,V. and Guben§ek,F. (1986) Bio. Chem.
Hoppe-Seyler 367, 919-923. 4. Kriiaj,I., Ritonja,A., Turk,D. and Gubensek,F. (1989) Biochem. Biophys. Acta 999, 198-202. 5. Bieber,A.L. personal communication. 6. Punger6ar,J., Kordis,D., Jerala,R., Trstenjak- Prebanda,M., Dolinar,M., Curin-Serbec,V., Komel,R. and Gubensek,F. (1989) Nucl. Acids Res. 17, 4367. 7. Kordis,D., Pungercar,J., 9tnukelj,B., Liang,N. and Gubensek,F. (1990) Nucl. Acids Res. 18, 4016. 8. van den Berg,C.J., Slotboom,A.J., Verheij,H.M. and de Haas,G.H. (1989) J. Cell. Biochem. 39, 379-390. 9. Kriiaj,I., Ritonja,A. and Gubensek,F. (1990) in preparation.
TGGACCCCCTTCAACTCTGAGACAAGGCTGCCAGCTGTCTGGATTCAGGAGG
-1
ATGAGGATTCTCTGGATAGTGGCCGTGTGCCTGATAGGCGTCGAGGGGAGCGTGATCGAA
60
M
R
I
L
W
I
V
A
V
C
L
I
G
V
E
G
S
V
I
20
E
TTTGGGAAGATGATCCAGGAGGAGACGGACAAAAACCCTCTTACCTCCTACTCCTTTTAC F
G
K
M
I
Q
E
E
T
D
K
N
P
L
T
S
Y
S
F
Y
120 40
G
C
H
C
G
L
G
N
K
G
K
P
K
D
A
T
D
R
C
C
180 60
TTCGTGCATAGCTGCTGTTACGCGAAACTGTCCGACTGCAGTCCCAAAACGAACAGATAC F
V
H
S
C
C
Y
A
K
L
S
D
C
S
P
K
T
N
R
GAATACCACCGGGAGAACGGGGCTATCGTCTGTGGAAGCAGCACCCCGTGCAAGAAACAG E
Y
H
R
E
N
G
A
I
V
C
G
S
S
T
P
C
K
K
240
80
Y
Q
300 100
360 I
C
E
C
D
R
A
A
A
I
C
F
R
E
N
L
K
T
Y
N
AAGAAATATAAGGTGTACCTGAGGTTTAAATGCAAGGGGGTTTCAGAGAAATGCTAAGTC K
K
Y
K
V
Y
L
R
F
K
C
K
G
V
S
E
K
C
*
120
420
138
TCTGCAGGCCGGGGAAAAACCCTCCAATTACACAATCGTAGTTGTGTTACTCTATTATTC 480 TGAATGCAATACTGAGCAATAAACAGGTGTCAGCTCTACACTCAAAAAAAAAAAAAAAAAA 541
*Permanent address: Laboratory of Pharmacology, Cancer Institute, Medical College, PO Box 530027, Nanning, China
