Different Research Approaches in Unraveling the Venom Proteome of Naja ashei

Abstract

The dynamic development of venomics in recent years has resulted in a significant increase in publicly available proteomic data. The information contained therein is often used for comparisons between different datasets and to draw biological conclusions therefrom. In this article, we aimed to show the possible differences that can arise, in the final results of the proteomic experiment, while using different research workflows. We applied two software solutions (PeptideShaker and MaxQuant) to process data from shotgun LC-MS/MS analysis of Naja ashei venom and collate it with the previous report concerning this species. We were able to provide new information regarding the protein composition of this venom but also present the qualitative and quantitative limitations of currently used proteomic methods. Moreover, we reported a rapid and straightforward technique for the separation of the fraction of proteins from the three-finger toxin family. Our results underline the necessary caution in the interpretation of data based on a comparative analysis of data derived from different studies.

Effects of 3FTx Protein Fraction from Naja ashei Venom on the Model and Native Membranes: Recognition and Implications for the Mechanisms of Toxicity

“…Protein concentration in each fraction was measured with Pierce™ BCA Protein Assay Kit (Thermo Scientific, Waltham, MA, USA) according to the manufacturer’s instruction. A detailed description of the procedure of the LC-MS/MS shotgun experiment with data analysis can be found in [ 8 ]. Briefly, for trypsin proteolysis, each sample contained 4.3 μg of proteins in 25 uL of 100 mM ammonium bicarbonate pH 8.…”

“…Neurotoxins (NT), cardiotoxins (or cytotoxins, CT), and phospholipase A 2 (PLA 2 ) are the three main groups of components that are responsible for the toxicity of Elapidae snake venoms [ 3 , 4 ], wherein NTs and CTs belong to the family of non-enzymatic proteins named three-finger toxins (3FTxs) [ 5 , 6 ]. 3FTxs are the most common toxins found in Elapidae venoms, including Naja ashei venom [ 7 , 8 ]. The species of this snake was designated in 2007 [ 9 ], which means that the structural and biochemical analysis of its venom’s effects on bioactive components on human cells has not been fully analyzed.…”

“…Additionally, unlike NT, CTs are amphiphilic and can cause cytotoxic effects to variety of cells. Some representatives of the CTx group occur in Naja ashei venom, e.g., CTxM1, CTxM4, and CTxM5 isoforms [ 7 , 8 ]. Different research studies indicate that this group of proteins has the ability to interact with lipid membranes.…”

Development of an Inhibition Enzyme-Linked Immunosorbent Assay (ELISA) Prototype for Detecting Cytotoxic Three-Finger Toxins (3FTxs) in African Spitting Cobra Venoms

“…N. ashei , also called the large brown spitting cobra was in 2007 considered a distinct species and classified accordingly [ 4 ]. As is the case with the other African spitting cobras, the venom of N. ashei is known to induce mainly cytotoxic effects caused by the predominant presence of 3FTx proteins and phospholipases A 2 [ 5 , 6 ]. N. ashei is a medically important snake species in Kenya, Ethiopia, Uganda, Somalia, and Tanzania, and thus, is implicated in snakebites and associated morbidity, mortality, and disability [ 7 ].…”

“…For instance, 3FTx proteins were found to constitute 73.3% of the total venom load of N. nigricollis [ 8 ]. Similarly, an analysis of the venom proteome of N. ashei revealed that 3FTx proteins accounted for 60–80% [ 5 , 6 ]. Three-finger toxins are non-enzyme-based polypeptides present in the venoms of elapids and colubrids [ 9 , 10 ], and are very rare in the venoms of vipers [ 11 ].…”