Isolation and Characterization of a Serine Protease, Ba III-4, from Peruvian Bothrops atrox venom

Ponce-Soto, Luis Alberto; Bonfim, V. L.; Novello, José Camillo; Oviedo, Ronald Demetrio Navarro; Chocas, Armando Yarlequé; Marangoni, Sérgio

doi:10.1007/s10930-007-9078-z

Cited by 18 publications

(23 citation statements)

References 24 publications

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“…Proteolytic activity was assayed according to Gutiérrez et al (2008) and Ponce et al (2007) [69,70] with modifications. Briefly, 100 µg of venom protein and 500 µL of 2% ( w/v ) casein (Sigma-Aldrich) were diluted in 0.1 mM Tris–HCl (pH 7.4) and 0.15 M NaCl buffer.…”

Section: Methodsmentioning

confidence: 99%

Snake Venom Hemotoxic Enzymes: Biochemical Comparison between Crotalus Species from Central Mexico

et al. 2019

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Snakebite envenoming is a serious medical problem in different areas of the world. In Latin America, the major prevalence is due to snakes of the family Viperidae, where rattlesnakes (Crotalus) are included. They produce hemotoxic venom which causes bleeding, tissue degradation and necrosis. Each venom has several enzymatic activities, producing different effects in the envenoming, doing its clinical effects difficult to study. Comparison between venom molecules is also difficult when different techniques are used, and therefore, their identification/characterization using the same methodology is necessary. In this work, a general biochemical characterization in snake venom of serine proteases (SVSP), phospholipases A2 (PLA2), metalloproteases (SVMP) and hyaluronidases (SVH) of Crotalus aquilus (Ca), Crotalus polystictus (Cp) and Crotalus molossus nigrescens (Cmn) was done. Differences in protein pattern, enzyme content and enzymatic activities were observed. All the venoms showed high PLA2 activity, high molecular weight SVSP, and a wide variety of SVMP and SVH forms. Ca and Cp showed the highest enzymatic activities of SVMP and SVSP trypsin-like and chymotrypsin-like, whereas Cmn showed the highest SVH and similar PLA2 activity with Ca. All the venoms showed peptides with similar molecular weight to crotamine-like myotoxins. No previous biochemical characterization of C. aquilus has been reported and there are no previous analyses that include these four protein families in these Crotalus venoms.

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Section: Methodsmentioning

confidence: 99%

Snake Venom Hemotoxic Enzymes: Biochemical Comparison between Crotalus Species from Central Mexico

et al. 2019

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“…Analysis by SDS-PAGE showed that TLBbar, has one band with ∼ 28.5 kDa, being a protein constituted of a single polypeptide chain (Figure 1(b) insert). TLBm of Bothrops marajoensis [9], Ba III-4 of Bothrops atrox [7], and PA-BJ of Bothrops jararaca [30] are serine proteases from snake venom that also have structure monomeric. Figure 2 shows the mass of TLBbar determined by mass spectrometry (ESI positive ionisation).…”

Section: Discussionmentioning

confidence: 99%

“…Snake venom serine proteases belong to trypsin S1 family clan SA. To date, a large number of studies, including molecular cloning, have led to the isolation and identification of TLEs mainly from the venom of subfamily Viperinaeand Crotalinae [3][4][5][6][7][8][9].…”

Section: Introductionmentioning

confidence: 99%

Biochemical and Pharmacological Characterization of TLBbar, a New Serine Protease with Coagulant Activity fromBothrops barnettiSnake Venom

et al. 2013

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A thrombin-like enzyme named TLBbar was isolated from Bothrops barnetti snake venom and its biochemical and pharmacological characteristics were determined. TLBbar was purified using size exclusion chromatography and reverse phase HPLC, showing molecular mass of 28750.7 Da determined by mass spectrometry. TLBbar serine protease is basic (pI 7.4) and its structure shows similarity with other serine proteases of snake venom. Optimal proteolytic activity was at 37 ∘ C and pH 8; this activity was strongly inhibited by PMSF and Leupeptin, however; heparin, and soybean trypsin inhibitor (SBT-I) were ineffective. Kinetic studies on BApNA chromogenic substrate have revealed that TLBbar presents a Michaelis-Menten kinetics, with values of and max of 0.433 mM and 0.42 nmol/min, respectively. TLBbar showed high clotting activity upon bovine and human plasma, presenting IC of 125 and minimum dose coagulant (MDC) of 2.23 g/ L. TLBbar cleavages the A chain of bovine fibrinogen, with maximal efficiency at 30-40 ∘ C in the presence of calcium after two hours incubation; this fibronogenolityc activity was inhibited by PMSF and Leupeptin, confirming its classification in the group of serine proteases. In addition, TLBbar is capable of aggregating platelets in the same way that thrombin in concentrations of 2.5 g/ L.

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“…Anti-proteolytic activity. Proteolytic activity was determined using the synthetic chromogenic substrate Nα-benzoyl-DL-arginine p-nitroanilide (DL-BApNA), in 96-well plates, according to Ponce-Soto et al (14). BjuV or mixtures of BjuV and BFRE (1:1, 1:2, 1:3, w:w) were diluted with buffer (Tris-HCl, 10 mM, 10 mM CaCl 2 , 100 mM NaCl, pH 7.8) at 2 mg/mL.…”

Section: Methodsmentioning

confidence: 99%

Anti-ophidian activity of Bredemeyera floribunda Willd. (Polygalaceae) root extract on the local effects induced by Bothrops jararacussu venom

Alves

Ximenes

Jorge

et al. 2019

Braz J Med Biol Res

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Bredemeyera floribunda roots are popularly used to treat snakebites in the semiarid region of Northeast Brazil, and previous studies indicate the anti-ophidian actions of triterpenoid saponins found in its roots. To assess B. floribunda root extract (BFRE) activity against the effects of Bothrops jararacussu venom (BjuV), antiphospholipasic, antiproteolytic, antihemorrhagic, antinecrotic, and anti-edematogenic activities were investigated in mice. Phytochemical analysis revealed the presence of saponins, flavonoids, and sugars, with rutin and saccharose being the major constituents of BFRE. Acute toxicity was determined and BFRE was nontoxic to mice. Phospholipase A2 and proteolytic activities induced by BjuV were inhibited in vitro by BFRE at all concentrations tested herein. BFRE (150 mg/kg) inhibited paw edema induced by BjuV (50 µg/animal), reducing total edema calculated by area under the curve, but carrageenan-induced paw edema was unchanged. Hemorrhagic and necrotizing actions of BjuV (50 µg/animal) were considerably decreased by BFRE treatment. Thus, BFRE blocked the toxic actions of B. jararacussu venom despite having no anti-inflammatory activity, which points to a direct inhibition of venom’s toxins, as demonstrated in the in vitro assays. The larger amounts of rutin found in BFRE may play a role in this inhibition, since 3′,4′-OH flavonoids are known inhibitors of phospholipases A2.

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Isolation and Characterization of a Serine Protease, Ba III-4, from Peruvian Bothrops atrox venom

Cited by 18 publications

References 24 publications

Snake Venom Hemotoxic Enzymes: Biochemical Comparison between Crotalus Species from Central Mexico

Snake Venom Hemotoxic Enzymes: Biochemical Comparison between Crotalus Species from Central Mexico

Biochemical and Pharmacological Characterization of TLBbar, a New Serine Protease with Coagulant Activity fromBothrops barnettiSnake Venom

Anti-ophidian activity of Bredemeyera floribunda Willd. (Polygalaceae) root extract on the local effects induced by Bothrops jararacussu venom

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