TY - JOUR
T1 - Biochemical and biological characterization of a PLA2 from crotoxin complex of Crotalus durissus cumanensis
AU - Pereañez, Jaime Andrés
AU - Núñez, Vitelbina
AU - Huancahuire-Vega, Salomón
AU - Marangoni, Sergio
AU - Ponce-Soto, Luis Alberto
N1 - Funding Information:
The authors thank Paulo A. Baldasso for general technical help. This project was supported by Universidad de Antioquia, COLCIENCIAS (project 393-2006) and CYTED (project 206AC0281). This study was performed as partial requirement for the PhD degree of Jaime Andrés Pereañez at Universidad de Antioquia.
PY - 2009/4
Y1 - 2009/4
N2 - A new PLA2 (Cdcum6) from crotoxin complex of Colombian Crotalus durissus cumanensis rattlesnake was purified using molecular exclusion chromatography and RP-HPLC. The molecular mass of Cdcum6 was determined by SDS-PAGE ∼14 KDa and confirmed by MALDI-TOF (14321.98 Da). The enzyme showed Km 6.0 mM, Vmax 3.44 nmol/min, optimum pH was 8.0 and temperature was between 30 and 45 °C, and it had a strict requirement of Ca2+ for its activity. The N-terminal sequence of PLA2 was SLVQF EKMIK EVAGK NGVPWY. Comparison of amino acid sequence data with other PLA2 from South American Crotalus durissus rattlesnakes showed that Cdcum6 shares the highest sequence identity with Cdr13 an isoform PLA2 from Crotalus durissus ruruima, nevertheless, Cdcum6 showed high content of basic and hydrophobic amino acids. In mice, Cdcum6 presented higher LD50 than crotoxin complex from C. d. cumanensis. Additionally, Cdcum6 induced a conspicuous local myotoxic effect and moderate footpad edema; in vitro, it was antigoagulant in doses as low as 0.5 μg/ml, and it was not cytotoxic on myoblast but Cdcum6 was able to lyse myotubes.
AB - A new PLA2 (Cdcum6) from crotoxin complex of Colombian Crotalus durissus cumanensis rattlesnake was purified using molecular exclusion chromatography and RP-HPLC. The molecular mass of Cdcum6 was determined by SDS-PAGE ∼14 KDa and confirmed by MALDI-TOF (14321.98 Da). The enzyme showed Km 6.0 mM, Vmax 3.44 nmol/min, optimum pH was 8.0 and temperature was between 30 and 45 °C, and it had a strict requirement of Ca2+ for its activity. The N-terminal sequence of PLA2 was SLVQF EKMIK EVAGK NGVPWY. Comparison of amino acid sequence data with other PLA2 from South American Crotalus durissus rattlesnakes showed that Cdcum6 shares the highest sequence identity with Cdr13 an isoform PLA2 from Crotalus durissus ruruima, nevertheless, Cdcum6 showed high content of basic and hydrophobic amino acids. In mice, Cdcum6 presented higher LD50 than crotoxin complex from C. d. cumanensis. Additionally, Cdcum6 induced a conspicuous local myotoxic effect and moderate footpad edema; in vitro, it was antigoagulant in doses as low as 0.5 μg/ml, and it was not cytotoxic on myoblast but Cdcum6 was able to lyse myotubes.
KW - Crotalus durissus cumanensis
KW - Crotoxin
KW - Myotoxin
KW - Neurotoxin
KW - Phospholipase A
UR - http://www.scopus.com/inward/record.url?scp=61649100259&partnerID=8YFLogxK
U2 - 10.1016/j.toxicon.2009.01.021
DO - 10.1016/j.toxicon.2009.01.021
M3 - Article
C2 - 19673100
AN - SCOPUS:61649100259
SN - 0041-0101
VL - 53
SP - 534
EP - 542
JO - Toxicon
JF - Toxicon
IS - 5
ER -