Polyacrylamide Gel Electrophoresis as a Tool for the Taxonomic Identification of Snakes From the Elapidae and Viperidae Families

Soares, Andreimar Martins; Pedrosa, L. H. Anzaloni; Fontes, M.R.M.; Silva, R. J. Da; Giglio, José Roberto

doi:10.1590/s0104-79301998000200005

Cited by 18 publications

(6 citation statements)

References 5 publications

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“…Lately, 2DE has been the preferred technique used by several researchers to decipher the complexity of venoms due to its capability of separating a mixture of proteins based on two parameters, pI and MW (15)(16)(17)(18)(19). Owing to the close proximity of molecular weights among toxins bearing different net charges, SDS-PAGE was not able to provide efficient differentiation (6). Other than working directly on venom, the use of 2DE has also been relevant on serum of a person bitten by rattlesnake, revealing qualitative and quantitative changes in plasma proteins which are obscure in 1D (20).…”

Section: Discussionmentioning

confidence: 99%

Protein profile analysis of Malaysian snake venoms by two-dimensional gel electrophoresis

Vejayan¹,

Yee²,

Ponnudurai³

et al. 2010

J. Venom. Anim. Toxins incl. Trop. Dis

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Snake venoms comprise a highly complex mixture of proteins, which requires for their characterization the use of versatile two-dimensional electrophoresis techniques. In the present study, venoms obtained from eight snakes (Ophiophagus hannah, Naja kaouthia, Naja sumatrana, Bungarus fasciatus, Trimeresurus sumatranus, Tropidolaemus wagleri, Enhydrina schistosa and Calloselasma rhodostoma) commonly found in Malaysia were separated based on two independent properties, isoelectric point (pI) and molecular weight (MW). Many differences in snake venoms at the inter-family, inter-subfamily, inter-genus and inter-species levels were revealed. Notably, proteins from individuals of the Viperidae family -Trimeresurus sumatranus, Tropidolaemus wagleri and Calloselasma rhodostoma -were found to be numerous and scattered by the two-dimensional gel electrophoresis (2DE) specifically in regions between 37 and 100 kDa compared to the Elapidae venom proteins. The latter were clustered at the basic and lower molecular mass region (less than 20 kDa). Trains of spots were commonly observed, indicating that these proteins may be derived from post-translational modifications. Ophiophagus hannah (Elapidae) revealed a great amount of protein spots in the higher molecular mass range when compared to Enhydrina schistosa, Naja kaouthia, Naja sumatrana and Bungarus fasciatus. Overall 2DE showed large differences in the venom profile of each species, which might be employed as an ancillary tool to the identification of venomous snake species.

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Section: Discussionmentioning

confidence: 99%

Protein profile analysis of Malaysian snake venoms by two-dimensional gel electrophoresis

Vejayan¹,

Yee²,

Ponnudurai³

et al. 2010

J. Venom. Anim. Toxins incl. Trop. Dis

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“…The fractionation and purification of different components of scorpion venom are mainly performed by chromatographic methods [21]. Moreover, the peculiar species-specific chromatographic profiles of venoms may also be used as an auxiliary tool in the taxonomy of scorpions [22][23][24][25][26]. In this study, we present the chromatographic profiles of three dreaded scorpion species from Saudi Arabia.…”

Section: Introductionmentioning

confidence: 99%

Rapid profiling of crude scorpion venom using liquid chromatography and its relevance to species identification

Asmari

Khan

Manthiri

2012

Acta Chromatographica

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Summary. We present the comprehensive chromatographic profiles of three scorpion species, Androctonus crassicauda, Androctonus bicolor, and Leiurus quinquestriatus, commonly inhabited to Middle East regions. Their venoms were milked by electrical stimulation, diluted with distilled water, properly mixed and centrifuged to separate the mucus from venom. The clear supernatant was filtered and the protein concentration was determined. Pre-diluted venoms were chromatographed on FPLC (fast protein liquid chromatography) and RP-HPLC (reversed-phase high-performance liquid chromatography) and the fractions were collected for molecular weight determination. Both techniques have resulted clearly distinguishable chromatographic patterns that can be used for identification of scorpion species and having a quick indication of venom toxicity.

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“…One-dimensional SDS-PAGE has limited differentiation efficiency for crude venom proteins, owing to the overlapping of protein bands with similar molecular weights (19). Protein spots on the 2DE profile are more specific and more definite indications of the presence of proteins in a particular sample.…”

Section: Discussionmentioning

confidence: 99%

The concept of two-dimensional electrophoresis-guided purification proven by isolation of rhodocetin from Calloselasma rhodostoma (Malayan pit viper)

Tang¹,

Vejayan²,

Ibrahim³

2011

J. Venom. Anim. Toxins incl. Trop. Dis

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Two-dimensional gel electrophoresis (2DE) is an important tool for investigating the complexity of snake venom proteomes. Apart from applications based on whole proteome analysis, we suggest that 2DE can be used as an assay to guide the progress of protein purification. The aim of this study was to prove the feasibility of this concept by using it to purify rhodocetin from Calloselasma rhodostoma venom. Rhodocetin (α subunit) spot on the 2DE profile of C. rhodostoma venom was first identified and confirmed by mass spectrometry, with a molecular mass of 16 kDa and calculated pI of 5.16. Rhodocetin was subsequently purified by successive anion-exchange and gel filtration chromatography. Every peak from both chromatography profiles was collected and tested on 2DE. The presence of rhodocetin (α subunit) spot in the 2DE profile of the peak DP2 indicated the presence of the protein. The purified compound was used to spike the crude venom. A spiked spot with a 1.6-fold increase in intensity was observed and its position matched to that of rhodocetin (α subunit) on the 2DE profile. Together, these spots confirmed the identity of the purified compound as rhodocetin. Hence, our results have demonstrated the effectiveness of the concept we now term 2DE-guided purification.

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Polyacrylamide Gel Electrophoresis as a Tool for the Taxonomic Identification of Snakes From the Elapidae and Viperidae Families

Cited by 18 publications

References 5 publications

Protein profile analysis of Malaysian snake venoms by two-dimensional gel electrophoresis

Protein profile analysis of Malaysian snake venoms by two-dimensional gel electrophoresis

Rapid profiling of crude scorpion venom using liquid chromatography and its relevance to species identification

The concept of two-dimensional electrophoresis-guided purification proven by isolation of rhodocetin from Calloselasma rhodostoma (Malayan pit viper)

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