Amino acid sequence and crystal structure of BaP1, a metalloproteinase from <i>Bothrops asper</i> snake venom that exerts multiple tissue‐damaging activities

Watanabe, Leandra; Shannon, John D.; Valente, Richard H.; Rucavado, Alexandra; Alape-Girón, Alberto; Kamiguti, Aura S.; Theakston, R.D.G.; Fox, Jay W.; Gutiérrez, José Marı́a; Arni, Raghuvir K.

doi:10.1110/ps.03102403

Cited by 124 publications

(79 citation statements)

References 48 publications

(111 reference statements)

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“…Similar observations were seen for other crystal structures of non-hemorrhagic metalloproteinases including the fibrinogenolases (TM-1, TM-2, and TM-3) from Taiwan Habu [58]. Indeed, these three non-hemorrhagic metalloproteinases contain a very robust basic field potential surface whereas acutolysin C and Bap1 from Bothrops asper, two weakly hemorrhagic metalloproteinases [53], show energy landscapes with mix polarities in that they contain a highly basic field potential landscape at the N-terminal region and an acidic field potential energy landscape near the catalytic site (data not shown). In further agreement with this view, the intensity of the negative field potential maps surrounding the Zn +2 binding domain and the Met turn for each of the C. s. scutulatus metalloproteinases correlated with hemorrhagic potency [32] in the following manner: atrolysin C> GP1> GP4> GP3> GP2> adamalysin II.…”

Section: Discussionsupporting

confidence: 83%

“…For instance, it has been reported that the interconnecting loop that follows the Met turn, which also contains the S'1-specificity pocket, is remarkably more flexible in non-hemorrhagic metalloproteinases compared to hemorrhagic metalloproteinases [52]. In agreement with this observation [52,53], the C. s. scutulatus metalloproteinases belonging to the non-hemorrhagic group of metalloproteinases (GP2) contained a much more dynamic S'1-pocket than the strongly or weakly hemorrhagic groups (GP1 and GP4) as measured by the standard deviations of the angular shift over time (GP1: ±4.04 • , GP2: ±6.72 • , GP3: ±6.73 • , GP4: ±4.90 • ).…”

Section: Discussionmentioning

confidence: 53%

“…Moreover, prior to our studies, several studies showed that the hemorrhagic potency of rattlesnake venom metalloproteinases correlated with the degree of flexibility of the interconnecting loops that precede or follow the Met turn [52,53]. For instance, it has been reported that the interconnecting loop that follows the Met turn, which also contains the S'1-specificity pocket, is remarkably more flexible in non-hemorrhagic metalloproteinases compared to hemorrhagic metalloproteinases [52].…”

Section: Discussionmentioning

confidence: 55%

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Molecular models of the Mojave rattlesnake (Crotalus scutulatus scutulatus) venom metalloproteinases reveal a structural basis for differences in hemorrhagic activities

Dagda

Gasanov²,

Zhang³

et al. 2014

J Biol Phys

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Rattlesnake venom can differ in composition and in metalloproteinase-associated activities. The molecular basis for this intra-species variation in Crotalus scutulatus scutulatus (Mojave rattlesnake) remains an enigma. To understand the molecular basis for intra-species variation of metalloproteinase-associated activities, we modeled the threedimensional structures of four metalloproteinases based on the amino acid sequence of four variations of the proteinase domain of the C. s. scutulatus metalloproteinase gene (GP1, GP2, GP3, and GP4). For comparative purposes, we modeled the atrolysin metalloproteinases of C. atrox as well. All molecular models shared the same topology. While the atrolysin metalloproteinase molecular models contained highly conserved substrate binding sites, the Mojave rattlesnake metalloproteinases showed higher structural divergence when superimposed onto each other. The highest structural divergence among the four C. s. scutulatus molecular models was located at the northern cleft wall and the S' 1 -pocket of the substrate binding site, molecular regions that modulate substrate selectivity. Molecular dynamics and field potential maps for each C. s. scutulatus metalloproteinase model demonstrated that the non-hemorrhagic metalloproteinases (GP2 and GP3) contain highly basic molecular and field potential surfaces while the hemorrhagic metalloproteinases GP1 and atrolysin C showed extensive acidic field potential maps and shallow but less dynamic active site pockets. Hence, differences in the spatial arrangement of the northern cleft wall, the S' 1 -pocket, and the physico-chemical environment surrounding the catalytic site contribute to differences in metalloproteinase activities in the Mojave rattlesnake. Our results provide a structural basis for variation of metalloproteinase-associated activities in the rattlesnake venom of the Mojave rattlesnake.

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Section: Discussionsupporting

confidence: 83%

Section: Discussionmentioning

confidence: 53%

Section: Discussionmentioning

confidence: 55%

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Molecular models of the Mojave rattlesnake (Crotalus scutulatus scutulatus) venom metalloproteinases reveal a structural basis for differences in hemorrhagic activities

Dagda

Gasanov²,

Zhang³

et al. 2014

J Biol Phys

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“…Este espectro foi analisado utilizando a técnica de bioinformática em MALDI TOF, com o programa Mascot Search Results (Matrix Science), o qual revelou score significativo (score de 80 e p<0.05) com a metaloprotease não hemorrágica BmooMPα-I, proveniente também da peçonha da serpente do gênero Bothrops moojeni (BERNADES et al, 2008 (WATANABE et al, 2003). Desta maneira, pode-se afirmar, de acordo com a classificação das metaloproteases apresentadas por FOX; SERRANO, (2010), que BthMP é uma metaloprotease da classe P-I.…”

Section: Resultsunclassified

Purificação e avaliação da interferência de BthMP (uma metaloprotease da peçonha de Bothrops moojeni) na coagulação sanguínea

et al. 2016

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In the present was work reported the purification of the metalloprotease BthMP, from the venom of Bothrops moojeni. For purification of the protease was used ion exchange chromatography (DEAE-Sepharose) and molecular exclusion , the product of these processes was a protein band with high purity, visualized on SDS-PAGE 14%, referred the BthMP. This protease when analyzed on MALDI-TOF revealed the molecular weight of the native form of 23.050 Da and 23.872 Da in the reduced form, and from the peptide fragments obtained by Peptide Mass Fingerprinting (PMF) in MS (MALDI-TOF/TOF) was observed high similarity with BmooMPα-I metalloprotease. In enzymatic terms, BthMP showed proteolytic activity on azocasein using PMSF and benzamidine, while this activity was inhibited in the presence of EDTA; 1,10-Phenanthroline and β-mercaptoethanol, it is therefore a zinc-dependent metalloprotease of the class P-I. To still with this purpose, we contemplate its enzymatic specificity of the Aα and Bβ chains of fibrinogen and also the consumption of fibrinogen in vivo. It was also found its action on the components of the coagulation cascade, due to prolongation of prothrombin time and partial thromboplastin time. Thus, the sharp fibrinogenolytic activity and the high consumption of fibrinogen in vivo are results that indicate the anticoagulant action of BthMP; furthermore, their ability to interfere with the coagulation cascade suggested that this protease is promising for future studies that might indicate a new antithrombotic agent model. K E Y W O R D SCoagulation cascade, Metalloproteinase, Venom snake R E S U M ONeste trabalho relatamos a purificação da metaloprotease BthMP, proveniente da peçonha da serpente Bothrops moojeni. Para a purificação desta protease, utilizaram-se os passos cromatográficos de troca iônica (DEAE-Sepharose) e de exclusão molecular (Sephadex G-75), sendo o produto desses processos uma banda proteica com elevado grau de pureza, visualizada em SDS-PAGE a 14%, denominada BthMP. Esta, por sua vez, quando analisada em MALDI-TOF revelou a massa molecular nativa de 23.050 Da e 23.872 Da na forma reduzida, e a partir dos fragmentos peptídicos obtidos por Peptide Mass Fingerprinting (PMF) em MS (MALDI-TOF/TOF) indicou alta similaridade com a metaloprotease BmooMPα-I. Em termos enzimáticos, BthMP mostrou atividade proteolítica sobre azocaseína e frente ao PMSF e benzamidina, enquanto que esta atividade foi inibida na presença de EDTA, 1,10-fenantrolina e β-mercaptoetanol, sendo portanto uma metaloprotease zinco dependente da classe P-I. Ainda com este propósito, verificou-se sua especificidade enzimática sobre as cadeias Aα e Bβ do fibrinogênio, e também o consumo de fibrinogênio in vivo. Foi constatado ainda sua ação em componentes da cascata de coagulação, devido ao prolongamento do Tempo de Protrombina (TP) e do Tempo de Tromboplastina Parcial ativada (TTPa). Desta forma, a acentuada atividade fibrinogenolítica e o alto consumo de fibrinogênio in vivo são resultados que indicam a ação anticoagulante da BthMP; al...

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“…A number of hemorrhagic SVMPs have been isolated from B. asper venom. The most abundant SVMP is the P-I enzyme BaP1 (Gutié rrez et al, 1995;Watanabe et al, 2003), although the most potent hemorrhagic enzymes are the high molecular mass P-III SVMPs BaH1 and BaH4 (Borkow et al, 1993;Franceschi et al, 2000). Moreover, the effectiveness of clodronate and doxycycline to inhibit coagulant activity of B. asper venom on plasma and defibrinogenating activity (in the case of doxycycline) can also be explained by the observation that SVMPs are the most important coagulant components in this venom (Rucavado et al, 2004).…”

Section: Discussionmentioning

confidence: 99%

Assessment of metalloproteinase inhibitors clodronate and doxycycline in the neutralization of hemorrhage and coagulopathy induced by Bothrops asper snake venom

Rucavado

Henríquez

García

et al. 2008

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a b s t r a c tSnake venom metalloproteinases (SVMPs) play a prominent role in the local and systemic manifestations of viperid snakebite envenomations. Thus, the possibility of using metalloproteinase inhibitors in the treatment of these envenomations is a promising therapeutic alternative. This study assessed the ability of two metalloproteinase inhibitors, the biphosphonate clodronate and the tetracycline doxycycline, to inhibit proteolytic, hemorrhagic, coagulant and defibrinogenating effects of Bothrops asper venom. Both compounds were able to inhibit these activities, at concentrations in the mM range, when incubated with venom prior to testing. However, when inhibition of hemorrhage was assessed in assays involving independent injection of venom and drug, inhibition was poor, even when these compounds were injected immediately after envenomation. These findings support the concept that the effectiveness of compounds, such as clodronate and doxycycline, whose inhibitory action on SVMPs is based on zinc chelation alone, is limited, and stress the view that more specific molecules are required for an effective inhibition of SVMPs in vivo.

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Amino acid sequence and crystal structure of BaP1, a metalloproteinase from Bothrops asper snake venom that exerts multiple tissue‐damaging activities

Cited by 124 publications

References 48 publications

Molecular models of the Mojave rattlesnake (Crotalus scutulatus scutulatus) venom metalloproteinases reveal a structural basis for differences in hemorrhagic activities

Molecular models of the Mojave rattlesnake (Crotalus scutulatus scutulatus) venom metalloproteinases reveal a structural basis for differences in hemorrhagic activities

Purificação e avaliação da interferência de BthMP (uma metaloprotease da peçonha de Bothrops moojeni) na coagulação sanguínea

Assessment of metalloproteinase inhibitors clodronate and doxycycline in the neutralization of hemorrhage and coagulopathy induced by Bothrops asper snake venom

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