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The scorpion of Androctinus crassicauda is one of the Buthida family members whose neurotoxic venom is deadly to humans. Enzymes are abundant components of animal venom that play roles in local and systemic symptoms following envenoming. The venom of this scorpions contains various pathological enzymes, among which studies have reported the toxic effects of venom's hyaluronidase and phospholipase A2 enzymes. This study aimed to measure the activity of venom enzymes of hyaluronidase and phospholipase A2 from scorpion of A. crassicauda and its fractions. For this purpose, the venom from scorpion of A. crassicauda were collected and lyophilized. The whole venom was fractionated on a gel filtration (Sephadex G-50) column. Then protein concentration, hyaluronidase and phospholipase A2 activity of the crude venom and its fractions were determined. Gel filtration on Sephadex G-50 and SDS-PAGE electrophoresis of A. crassicauda crude venom revealed four protein picks and 15 protein bands with the majority of molecular masses between 10.0 and 15.0 kDa, respectively. The supernatant obtained from crude venom clarification showed protein content 71.80%, high hyaluronidase (92.40%) and low PLA2 (302±2.1 mg/U) enzyme activity. Fraction 1 showed highest hyaluronidase and phospholipase A2 activities. We found that pH 5 is the optimum pH for hyaluronidase activity and phospholipase A2 activity increases with increasing incubation time. In conclusion, venom of A. crassicauda displayed the hyaluronidase and phospholipase A2 enzymes activity, which were separated by gel filtration chromatography in fraction I. Such information is important both for predicting the biological activity of venom components and useful for developing effective antivenoms based on these venom components. [ABSTRACT FROM AUTHOR]

Scorpions are the oldest terrestrial fauna with an ancient evolutionary history. To study the origins of scorpions, we conducted a molecular phylogenetic study of the arthropod using slowly evolving enolase sequences. In this study, we used deep RNA-seq technology to identify the enolase sequences of Androctonus crassicauda and Hemiscorpius lepturus scorpions. The scorpion's enolase sequences were in silico analyzed to predict the protein structures, physicochemical characterization, and exon-intron organization. The enolase of the scorpion was found to be a protein of 433 amino acids with a molecular mass of ∼47 kDa and an acidic isoelectric point. Here, we reported the existence of 8 exons and an inner GC/AG intron for scorpion enolase. Using homology modeling, the enolase of the scorpions was found to be very conservative during the evolution process. Furthermore, our phylogenetic analysis indicated the independent evolution of the Buthidae family from other scorpions and also the sister group relationship of Hemiscorpionidae with the Caraboctonidae families. The scorpions' enolase sequences demonstrated a close fit to those of spiders, followed by mite counterparts. This strongly supports our phylogeny analysis, which indicates a sister-group relationship between scorpions and spiders and their common ancestors with mites. This study provided support for the pairing of scorpions with spiders, mites, etc. using enolase as a nuclear gene. Our molecular clock analyzes of the arthropod show that the most recent common ancestor of arachnids and insects were diversified in the Cryogenian period of Neoproterozoic era, earlier than dates reported from fossil records. [ABSTRACT FROM AUTHOR]

Venom hyaluronidase is an enzyme belonging to the glycoside hydrolase family that plays a critical role in systemic envenomation by spreading toxins in tissue and destruction of the extracellular matrix. The roles of venom hyaluronidase during envenomation have been widely studied, but the identification of its variants and isoforms is still under investigation. In this study, we developed a filtering method to identify the exon-intron pattern, alternative splicing events and isoforms of hyaluronidase in A. crassicauda and H. lepturus scorpions using the RNA-seq technique. Furthermore, in silico analysis was performed to identify and characterize the hyaluronidases. The most important findings were that the scorpion hyaluronidase gene contains 5 exons, 4 introns and undergo to alternative splicing events. In A. crassicauda and H. lepturus datasets, a sequence denominated AcHase-1 and three denominated HLHase1, HLHase2 and HLHase3 were identified as hyaluronidase variants respectively, which were found to have multiple isoforms that differed in the coding, non-coding or untranslated regions. The results showed that exon skipping, intron retention and alternative 3' splice site led to dysregulation of gene expression. To clarify the evolutionary history of scorpions based on hyaluronidase molecular phylogenetic studies, we used the phylogenetic tree to deduce the origins of this protein in different species of scorpions, spiders and bees. Hyaluronidase gene performed well for divergences and accurately separated closely related species. Our work provides insights into the diversity of Hase proteins during scorpion evolution and may direct further studies consisting of synthesis, purification and recombinant production of Hase proteins. [ABSTRACT FROM AUTHOR]