TY - JOUR
T1 - Biological and biochemical characterization of two new PLA2 isoforms Cdc-9 and Cdc-10 from Crotalus durissus cumanensis snake venom
AU - Romero-Vargas, Frey Francisco
AU - Ponce-Soto, Luis Alberto
AU - Martins-de-Souza, Daniel
AU - Marangoni, Sergio
N1 - Funding Information:
The authors thank Marino Forner for general technical help. This work was supported by CAPES and is part of Ms Sc thesis by Frey Francisco Romero-Vargas.
PY - 2010/1
Y1 - 2010/1
N2 - This work reports the purification, biological characterization and amino acid sequence of two new basic PLA2 isoforms, Cdc-9 and Cdc-10, purified from the Crotalus durissus cumanensis venom by one step analytical chromatography reverse phase HPLC. The molecular masses of the PLA2 were 14,175 ± 2.7 Da for Cdc-9 and 14,228 ± 3.5 Da for Cdc-10 both deduced by primary structure and confirmed by MALDI-TOF. The isoforms presented an amino acid sequence of 122 amino acid residues, being Cdc-9: SLVQFNKMIK FETRKSGLPF YAAYGCYCGW GGQRPKDATD RCCFVHDCCY GKVAKCNTKW DIYSYSLKSG YITCGKGTWC KEQICECDRV AAECLRRSLS TYKNEYMFYP DSRCREPPEY TC with pI value of 8.25 and Cdc-10: SLLQFNKMIK FETRKSGVPF YAAYGCYCGW GGRRPKDPTD RCCFVHDCCY GKLTKCNTKW DIYSYSLKSG YITCGKGTWC KEQICECDRV AAECLRRSLN TYKNEYMFYP DSRCRGPPEY TC with a pI value of 8.46, showing highly conserved Ca2+-binding and catalytic sites. The PLA2 activity decreased when the isoforms Cdc-9 and Cdc-10 were incubated with 4-bromophenacyl bromide (p-BPB), anhydrous acetic acid and p-nitrobenzene sulfonyl fluoride (NBSF) when compared with the activity of both native isoforms. In mice, the PLA2 isoforms Cdc-9 and Cdc-10 induced myonecrosis and edema. Myotoxic and edema activities were reduced after treatment of the isoforms with p-BPB; acetylation of the lysine residues and the treatment of PLA2 with NBSF have also induced edema reduction. However, p-BPB strongly diminishes the local and systemic myotoxic effects.
AB - This work reports the purification, biological characterization and amino acid sequence of two new basic PLA2 isoforms, Cdc-9 and Cdc-10, purified from the Crotalus durissus cumanensis venom by one step analytical chromatography reverse phase HPLC. The molecular masses of the PLA2 were 14,175 ± 2.7 Da for Cdc-9 and 14,228 ± 3.5 Da for Cdc-10 both deduced by primary structure and confirmed by MALDI-TOF. The isoforms presented an amino acid sequence of 122 amino acid residues, being Cdc-9: SLVQFNKMIK FETRKSGLPF YAAYGCYCGW GGQRPKDATD RCCFVHDCCY GKVAKCNTKW DIYSYSLKSG YITCGKGTWC KEQICECDRV AAECLRRSLS TYKNEYMFYP DSRCREPPEY TC with pI value of 8.25 and Cdc-10: SLLQFNKMIK FETRKSGVPF YAAYGCYCGW GGRRPKDPTD RCCFVHDCCY GKLTKCNTKW DIYSYSLKSG YITCGKGTWC KEQICECDRV AAECLRRSLN TYKNEYMFYP DSRCRGPPEY TC with a pI value of 8.46, showing highly conserved Ca2+-binding and catalytic sites. The PLA2 activity decreased when the isoforms Cdc-9 and Cdc-10 were incubated with 4-bromophenacyl bromide (p-BPB), anhydrous acetic acid and p-nitrobenzene sulfonyl fluoride (NBSF) when compared with the activity of both native isoforms. In mice, the PLA2 isoforms Cdc-9 and Cdc-10 induced myonecrosis and edema. Myotoxic and edema activities were reduced after treatment of the isoforms with p-BPB; acetylation of the lysine residues and the treatment of PLA2 with NBSF have also induced edema reduction. However, p-BPB strongly diminishes the local and systemic myotoxic effects.
KW - Crotalus durissus cumanensis
KW - Myotoxins
KW - RP-HPLC
KW - Snake venom
UR - http://www.scopus.com/inward/record.url?scp=71649092384&partnerID=8YFLogxK
U2 - 10.1016/j.cbpc.2009.08.011
DO - 10.1016/j.cbpc.2009.08.011
M3 - Article
C2 - 19747981
AN - SCOPUS:71649092384
SN - 1532-0456
VL - 151
SP - 66
EP - 74
JO - Comparative Biochemistry and Physiology - C Toxicology and Pharmacology
JF - Comparative Biochemistry and Physiology - C Toxicology and Pharmacology
IS - 1
ER -