ReviewOphidian envenomation strategies and the role of purines
Introduction
Recently, in the course of analyzing enzymatic compositional patterns relative to prey preference among coral snake taxa (Jorge da Silva and Aird, 2001) it was difficult to explain the ubiquitous nature of some relatively non-toxic venom constituents. The nearly universal occurrence of some enzymes suggested a central role in envenomation, but no satisfactory explanation has ever been given for their presence in venoms. As recently as 1999, Cousin and Bon (1999) noted that, ‘snake venoms are rich in proteins and enzymes whose functions are unknown’, citing nerve growth factor (NGF), L-amino acid oxidase (LAO) and phosphodiesterase (PDE) as examples. Numerous other enzymes and non-enzymatic toxins could also have been added to this short list.
At the same time, research in the Laboratório de Toxinas Naturais confirmed the earlier observation of Francis et al. (1997) that venom of the Brazilian coral snake, Micrurus frontalis, contains significant quantities of guanosine. Preliminary evidence suggested that a significant portion of the nucleoside is not free in solution, but that it is apparently tightly bound to postsynaptic neurotoxins and perhaps to phospholipases and other compounds as well. Intrigued by these findings I set out in search of an explanation for the role of purines in venoms.
Micrurus frontalis venom is far from being unique with regard to its purine content. The first such report appeared nearly 50 years ago when Fischer and Dörfel (1954) found adenosine in venoms of the puff adder (Bitis arietans), a large viperid, and of the eastern green mamba (Dendroaspis angusticeps), an elapid, and suggested that this compound might contribute to the hypotension promoted by these venoms. Doery (1956) reported the presence of adenosine, adenosine 3′-phosphate and ‘guanine compounds’ in venoms of several dissimilar elapids, Acanthophis antarcticus, Notechis scutatus, Dendroaspis angusticeps and Denisonia superba (Austrelaps superbus), and also in that of Bitis arietans. Doery (1957) later isolated guanosine, inosine and hypoxanthine from Notechis scutatus venom. Eight years passed before Wei and Lee (1965) reported that guanosine constitutes slightly over 1% of crude Bungarus multicinctus venom, but found no evidence that it influenced blood pressure in cats. Lo and Chen (1966) found adenosine, guanosine and inosine in the venom of the Chinese cobra, Naja atra. Lin and Lee (1971) suggested that nucleoside contamination might explain the detection of carbohydrate (ribose or deoxyribose) in many apparently pure elapid neurotoxins. Eterovic et al. (1975) reported that Bungarus multicinctus venom contains large amounts of free guanosine which accounted for 10% of the 280 nm absorbance of the crude venom. This material did not bind to a CM Sephadex column equilibrated in 50 mM ammonium acetate (pH 7.0). Takasaki et al. (1991) also identified adenosine, guanosine, and inosine in venom of the long-glanded coral snake (Maticora bivirgata).
In the present paper I outline three fundamental ophidian envenomation strategies which appear to be employed by all advanced venomous snakes. I suggest that purines act as multifunctional toxins, exerting synchronous effects upon virtually all cell types, and I believe that exogenous and released endogenous purines play a central role in all three envenomation strategies, which previously has been almost entirely overlooked. I offer hypothetical explanations for the presence of many venom enzymes which has hitherto been unexplained. Direct experimental evidence from venom studies is still lacking for most of the hypotheses presented herein, but pharmacological research in other fields provides ample evidence to suggest that they are probably correct and that they are worthy of experimental verification.
Section snippets
Venom functions
Snake venoms are extremely complex mixtures of proteins, peptides, carbohydrates, lipids, metal ions and organic compounds, with proteins and peptides accounting for approximately 90% of the dry weight (Bieber, 1979). While snake venoms have an obvious role in self-defense, this is of relatively little importance with regard to venom composition. Their primary function is to immobilize and kill prey organisms (Karlsson, 1979). Venoms simultaneously commence digestion of the prey from within,
Nomenclature
Two groups of nitrogenous bases are of immense importance in biological systems. The structurally more simple pyrimidines are six-sided, heterocyclic, aromatic rings, containing two nitrogen and four carbon atoms. The most important naturally occurring pyrimidines include cytosine, uracil and thymine (Fig. 1). Purines, with which the present paper is primarily concerned, comprise a fused ring system consisting of the pyrimidine ring linked to a five-sided imidazole ring. Five purines are well
Known synergistic actions of snake venom constituents
Historically snake venom research has necessarily been quite reductionist because only by isolating and characterizing individual components has it been possible to begin to understand the complex pathological state induced by envenomation. Perhaps because the majority of venom research has been done by biochemists and pharmacologists, there has also been a tendency to ignore the fact that venoms are optimized to function in specific prey organisms as integrated assemblages of compounds, and
Hypotensive strategy
Most snake venoms employ a variety of means to induce rapid and profound hypotension, leading to circulatory shock, prey immobilization and death (Bjarnason et al., 1983). The hypotensive peptides and kininogenases discussed above are only two of the many mechanisms employed for this purpose (Fig. 5). Several other well known hypotensive mechanisms will be briefly summarized before a variety of previously unrecognized pharmacological strategies are discussed.
Paralytic strategy
As with the hypotensive strategy, there are many paralytic strategy elements that are well known, and it is not the intent of this paper to review these; however, an incomplete picture would be presented if some of the better known examples were not mentioned briefly.
Digestive strategy
Prey immobilization is the first priority in all envenomation strategies and all snake venom components depend upon the prey's circulatory system for distribution throughout prey tissues (Fig. 5, Fig. 7). Nonetheless prey digestion also commences at the instant of envenomation and undoubtedly continues beyond the prey's death until venom constituents are inactivated by prey protease inhibitors or proteases, or by the snake's digestive enzymes. Some venom constituents have digestive functions
Role of venom purines other than adenosine
As previously mentioned, venoms have also been reported to contain guanosine, inosine and hypoxanthine. What possible contributions do these make to the envenomation strategies of the snakes that employ them?
Purine interactions with other toxins and miscellaneous roles
Lin and Lee (1971) suggested that nucleoside contamination might explain the detection of carbohydrate (ribose or deoxyribose) in many apparently pure toxins. I suggest that many toxins bind adenosine, guanosine and possibly inosine and that these bound purines also play an essential role in envenomation. If this hypothesis proves correct, much of the pharmacological data gathered with such toxins to date is probably in error. Specifically, toxin research involving neurotransmitter release,
Supporting evidence from arthropod venoms
Snake venoms are not the only toxic animal secretions to employ purines. Nucleotides and nucleosides are also important components of various spider venoms and at least one ant venom. Chan et al. (1975) discovered ATP, ADP and AMP in venoms of the tarantulas (Theraphosidae), Dugesiella and Aphonopelma. ATP, the principal nucleotide constituent, was present at a concentration of approximately 55 mM in Dugesiella, and 100 mM in Aphonopelma. ATP acted synergistically with the necrotoxin, the major
Conclusions
- •
Snake envenomation employs three well integrated strategies: prey immobilization via hypotension, prey immobilization via paralysis, and prey digestion.
- •
Purines (especially adenosine, inosine and guanosine) are concluded to play a central role in envenomation by most advanced venomous snakes.
- •
Purines constitute the perfect multifunctional toxins, participating simultaneously in all three envenomation strategies. Because they are endogenous regulatory and homeostatic compounds in all vertebrates,
Acknowledgements
I express my gratitude to my wife, Yayoi, whose love and support made this research possible, and also to my parents, John S. and Laurel J. Aird, for photocopying several hundred publications at the National Library of Medicine and the National Institutes of Health Library, without which this study could not have been done in Brazil. I gratefully acknowledge the assistance of Dr Saad Lahlou, not only for extensive editorial criticisms of the manuscript, but for his expertise in interpreting the
References (817)
- et al.
The heparin binding site of human extracellular-superoxide dismutase
Arch. Biochem. Biophys.
(1992) - et al.
Oral L-arginine improves endothelium-dependent dilatation and reduces monocyte adhesion to endothelial cells in young men with coronary artery disease
Atherosclerosis
(1997) - et al.
Toxins from the venom of the green mamba Dendroaspis angusticeps that inhibit the binding of quinuclidinyl benzilate to muscarinic acetylcholine receptors
Biochim. Biophys. Acta
(1988) - et al.
Characterization and cytotoxicity of L-amino acid oxidase from the venom of king cobra (Ophiophagus hannah)
Int. J. Biochem. Cell Biol.
(1997) - et al.
Comparative studies on three rattlesnake toxins
Toxicon
(1985) - et al.
Comparative enzymatic composition of Brazilian coral snake (Micrurus) venoms
Comp. Biochem. Physiol.
(1991) - et al.
Sea snake Hydrophis cyanocinctus venom. II. Histopathological changes, induced by a myotoxic phospholipase A2 (PLA2-H1)
Toxicon
(2000) - et al.
Purification by HPLC anion-exchange chromatography and some properties of a kinin-releasing enzyme from the venom of Agkistrodon halys ussuriensis
Toxicon
(1994) - et al.
Cardiovascular effects elicited by central administration of physostigmine via M2 muscarinic receptors in conscious cats
Brain Res.
(1995) - et al.
Renal cortical necrosis following Bothrops jararaca and B. jararacussu snake bite
Toxicon
(1985)
The effects of protease inhibitor homologues from mamba snake venoms on autonomic neurotransmission
Toxicon
Effects of fasciculin 2, an anticholinesterase polypeptide from green mamba venom, on neuromuscular transmission in mouse diaphragm preparations
Neurosci. Lett.
Enkephalin-processing oligopeptidases in cobra venom: Inhibition by thiorphan and bestatin reveals co-operative actions
Toxicon
A thrombin-like enzyme from bushmaster (Lachesis muta stenophrys) venom
Toxicon
Induction of apoptosis by hemorrhagic snake venom in vascular endothelial cells
Biochem. Biophys. Res. Commun.
Hemorrhagic, fibrinogenolytic and edema-forming activities of the venom of the colubrid snake Philodryas olfersii (green snake)
Toxicon
Histological and ultrastructural changes of the kidney in renal failure after viper envenomation
Toxicon
Immunohistochemical colocalization of the α-subunit of neutrophil NADPH oxidase and ecto-5′-nucleotidase in kidney and liver
Kidney Int.
TAFI, or plasma procarboxypeptidase B, couples the coagulation and fibrinolytic cascades through the thrombin–thrombomodulin complex
J. Biol. Chem.
Comparative effects of a selective A2 receptor agonist, CGS 21680, and nitroprusside in vascular smooth muscle
Eur. J. Pharmacol.
Central effects of adenosine analogs on locomotor activity in mice and antagonism of caffeine
Brain Res.
Anticonvulsant effects of adenosine analogues on amygdala-kindled seizures in rats
Neurosci. Lett.
Adenosine A2a receptors in the nucleus accumbens mediate locomotor depression
Brain Res. Bull.
Interactions of exendin-(9-39) with the effects of glucagon-like peptide-1-(7–36) amide and of exendin-4 on arterial blood pressure and heart rate in rats
Regul. Pept.
Purification and characterisation of a myotoxic phospholipase A2 from Indian cobra (Naja naja naja) venom
Toxicon
Purification and characterization of a neurotoxic phospholipase A2 from Indian cobra (Naja naja naja) venom
Toxicon
Kallikrein-like enzymes from Crotalus atrox venom
J. Biol. Chem.
The regulation of vascular function by P2 receptors: multiple sites and multiple receptors
Trends Pharmacol. Sci.
P2 purinoceptors on vascular endothelial cells: physiological significance and transduction mechanisms
Trends Pharmacol. Sci.
A phospholipase A2 with anticoagulant activity I. Isolation from Vipera berus venom and properties
Biochim. Biophys. Acta
Effects of centrally or systemically injected glucagon-like peptide-1 (7–36) amide on release of neurohypophysial hormones and blood pressure in the rat
Regul. Pept.
A simple assay for ecto-5′-nucleotidase using intact pulmonary artery endothelial cells. Effect of endotoxin-induced cell injury
Biochem. Pharmacol.
The contribution of residues 192 and 193 to the specificity of snake venom serine proteinases
J. Biol. Chem.
Focal cerebral ischemia enhances glial expression of ecto-5′-nucleotidase
Brain Res.
Nitric oxide: a signal for ADP-ribosylation of proteins
Life Sci.
Interaction between nerve growth factor and lysophosphatidylserine on rat peritoneal mast cells
FEBS Lett.
Comparative action of various kininogenases on crude horse plasma substrates
Biochem. Pharmacol.
Characterization of muscarinic receptors mediating vasodilation in guinea-pig ileum submucosal arterioles by the use of computer-assisted videomicroscopy
Eur. J. Pharmacol.
Evidence for prejunctional M2 muscarinic receptors in pulmonary cholinergic nerves in the rat
Br. J. Pharmacol.
Characterization of an apoptosis-inducing factor in Habu snake venom as a glycyrrhizin (GL)-binding protein potently inhibited by GL in vitro
Biol. Pharm. Bull.
The role of adenosine in insulin-induced vasodilation
J. Cardiovasc. Pharmacol.
Acute renal failure after envenomation by the common brown snake
Med. J. Aust.
Oral L-arginine inhibits platelet aggregation but does not enhance endothelium-dependent dilation in healthy young men
J. Am. Coll. Cardiol.
Disparate effects of A1- and A2-receptor agonists on intrarenal blood flow
Am. J. Physiol.
Ecto-5′-nucleotidase activity in lymphocytes from healthy and leukemia patients
Adv. Exp. Med. Biol.
Dipeptidyl(amino)peptidase IV and aminopeptidase M metabolize circulating substance P in vivo
J. Pharmacol. Exp. Ther.
ADP-ribosylation of actin by clostridial toxins
J. Cell Biol.
ADP-ribosylation of actin
J. Muscle Res. Cell Motil.
Adenosine receptor-mediated bronchoconstriction and bronchial hyperresponsiveness in allergic rabbit model
Am. J. Physiol.
Afibrinogenemia following snake bite (Crotalus durissus terrificus)
Am. J. Trop. Med. Hyg.
Cited by (243)
Phytochemical characterization and phospholipase A<inf>2</inf> inhibitory effect of Vitex negundo L. root extracts
2024, Journal of EthnopharmacologyEthylene glycol and plantain peel fraction ripened with calcium carbide inhibit reproductive mediators in male rats
2023, Endocrine and Metabolic ScienceIsolation and characterization of the first phosphodiesterase (Bj-PDE) from the venom of Bothrops jararacussu snake
2023, International Journal of Biological MacromoleculesAnimal Toxins
2023, Haschek and Rousseaux's Handbook of Toxicologic Pathology, Volume 3: Environmental Toxicologic Pathology and Major Toxicant Classes