Dataset Information

Venom variation in hemostasis of the southern Pacific rattlesnake (Crotalus oreganus helleri): isolation of hellerase.

ABSTRACT: Envenomations by the southern Pacific rattlesnake (Crotalus oreganus helleri) are the most common snakebite accidents in southern California. Intraspecies venom variation may lead to unresponsiveness to antivenom therapy. Even in a known species, venom toxins are recognized as diverse in conformity with interpopulational, seasonal, ontogenetic and individual factors. Five venoms of individual C. oreganus helleri located in Riverside and San Bernardino counties of southern California were studied for their variation in their hemostatic activity. The results demonstrated that Riverside 2 and San Bernardino 1 venoms presented the highest lethal activity without hemorrhagic activity. In contrast, San Bernardino 2 and 3 venoms had the highest hemorrhagic and fibrinolytic activities with low lethal and coagulant activities. Riverside 1, Riverside 2 and San Bernardino 1 venoms presented a significant thrombin-like activity. San Bernardino 2 and 3 venoms presented an insignificant thrombin-like activity. In relation to the fibrinolytic activity, San Bernardino 3 venom was the most active on fibrin plates, which was in turn neutralized by metal chelating inhibitors. These results demonstrate the differences amongst C. oreganus helleri venoms from close localities. A metalloproteinase, hellerase, was purified by anionic and cationic exchange chromatographies from San Bernardino 3 venom. Hellerase exhibited the ability to break fibrin clots in vitro, which can be of biomedically importance in the treatment of heart attacks and strokes.

SUBMITTER: Salazar AM

PROVIDER: S-EPMC2706139 | biostudies-literature | 2009 Apr

REPOSITORIES: biostudies-literature

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Venom variation in hemostasis of the southern Pacific rattlesnake (Crotalus oreganus helleri): isolation of hellerase.

Salazar Ana Maria AM Guerrero Belsy B Cantu Bruno B Cantu Esteban E Rodríguez-Acosta Alexis A Pérez John C JC Galán Jacob A JA Tao Andy A Sánchez Elda E EE

Comparative biochemistry and physiology. Toxicology & pharmacology : CBP 20080905 3

Envenomations by the southern Pacific rattlesnake (Crotalus oreganus helleri) are the most common snakebite accidents in southern California. Intraspecies venom variation may lead to unresponsiveness to antivenom therapy. Even in a known species, venom toxins are recognized as diverse in conformity with interpopulational, seasonal, ontogenetic and individual factors. Five venoms of individual C. oreganus helleri located in Riverside and San Bernardino counties of southern California were studied ...[more]

PMID: 18804187

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Project description:As trophic adaptations, rattlesnake venoms can vary in composition depending on several intrinsic and extrinsic factors. Ontogenetic changes in venom composition have been documented for numerous species, but little is known of the potential age-related changes in many rattlesnake species found in México. In the current study, venom samples collected from adult and neonate Crotalus polystictus from Estado de México were subjected to enzymatic and electrophoretic analyses, toxicity assays (LD50), and MALDI-TOF mass spectrometry, and a pooled sample of adult venom was analyzed by shotgun proteomics. Electrophoretic profiles of adult males and females were quite similar, and only minor sex-based variation was noted. However, distinct differences were observed between venoms from adult females and their neonate offspring. Several prominent bands, including P-I and P-III snake venom metalloproteinases (SVMPs) and disintegrins (confirmed by MS/MS) were present in adult venoms and absent/greatly reduced in neonate venoms. Age-dependent differences in SVMP, kallikrein-like, phospholipase A₂ (PLA₂), and L-amino acid oxidase (LAAO) activity levels were confirmed by enzymatic activity assays, and like many other rattlesnake species, venoms from adult snakes have higher SVMP activity than neonate venoms. Conversely, PLA₂ activity was approximately 2.5 × greater in venoms from neonates, likely contributing to the increased toxicity (neonate venom LD50 = 4.5 μg/g) towards non-Swiss albino mice when compared to adult venoms (LD50 = 5.5 μg/g). Thrombin-like (TLE) and phosphodiesterase activities did not vary significantly with age. A significant effect of sex (between adult male and adult female venoms) was also observed for SVMP, TLE, and LAAO activities. Analysis of pooled adult venom by LC-MS/MS identified 14 toxin protein families, dominated by bradykinin-inhibitory peptides, SVMPs (P-I, P-II and P-III), disintegrins, PLA₂s, C-type-lectins, CRiSPs, serine proteinases, and LAAOs (96% of total venom proteins). Neonate and adult C. polystictus in this population consume almost exclusively mammals, suggesting that age-based differences in composition are related to physical differences in prey (e.g., surface-to-volume ratio differences) rather than taxonomic differences between prey. Venoms from adult C. polystictus fit a Type I pattern (high SVMP activity, lower toxicity), which is characteristic of many larger-bodied rattlesnakes of North America.

Project description:BACKGROUND: Snake venoms have significant impacts on human populations through the morbidity and mortality associated with snakebites and as sources of drugs, drug leads, and physiological research tools. Genes expressed by venom-gland tissue, including those encoding toxic proteins, have therefore been sequenced but only with relatively sparse coverage resulting from the low-throughput sequencing approaches available. High-throughput approaches based on 454 pyrosequencing have recently been applied to the study of snake venoms to give the most complete characterizations to date of the genes expressed in active venom glands, but such approaches are costly and still provide a far-from-complete characterization of the genes expressed during venom production. RESULTS: We describe the de novo assembly and analysis of the venom-gland transcriptome of an eastern diamondback rattlesnake (Crotalus adamanteus) based on 95,643,958 pairs of quality-filtered, 100-base-pair Illumina reads. We identified 123 unique, full-length toxin-coding sequences, which cluster into 78 groups with less than 1% nucleotide divergence, and 2,879 unique, full-length nontoxin coding sequences. The toxin sequences accounted for 35.4% of the total reads, and the nontoxin sequences for an additional 27.5%. The most highly expressed toxin was a small myotoxin related to crotamine, which accounted for 5.9% of the total reads. Snake-venom metalloproteinases accounted for the highest percentage of reads mapping to a toxin class (24.4%), followed by C-type lectins (22.2%) and serine proteinases (20.0%). The most diverse toxin classes were the C-type lectins (21 clusters), the snake-venom metalloproteinases (16 clusters), and the serine proteinases (14 clusters). The high-abundance nontoxin transcripts were predominantly those involved in protein folding and translation, consistent with the protein-secretory function of the tissue. CONCLUSIONS: We have provided the most complete characterization of the genes expressed in an active snake venom gland to date, producing insights into snakebite pathology and guidance for snakebite treatment for the largest rattlesnake species and arguably the most dangerous snake native to the United States of America, C. adamanteus. We have more than doubled the number of sequenced toxins for this species and created extensive genomic resources for snakes based entirely on de novo assembly of Illumina sequence data.

Dataset Information

Venom variation in hemostasis of the southern Pacific rattlesnake (Crotalus oreganus helleri): isolation of hellerase.

Publications

Venom variation in hemostasis of the southern Pacific rattlesnake (Crotalus oreganus helleri): isolation of hellerase.

Similar Datasets

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