Bucandin

Bucandin is a novel, presynaptic neurotoxin that comes from the venom of the Malayan Krait snake, which is also known as its species name, Bungarus candidus. It is unique in many ways, but most importantly in how it operates within the human body when it enters after the initial snake bite.

Size

When it comes to a protein, it can help to first understand the basics of it. A few notable pieces of information are the total structure weight of the protein, the atom count of the protein, and the residue count. The total structure weight of the protein tells us how large the protein is as it gives us an insight as to how heavy it is compared to other proteins. Bucandin, for example, has a total structure weight of 7293.41 g/mol. In relation to other proteins, this is a relatively small one. When looking at another protein, activated unliganded spinach rubisco (1aus), for example, Bucandin is dwarfed by it. The rubisco protein has a molecular weight of 269774.72 g/mol, which is nearly 37 times larger in weight than Bucandin. The atom count of a molecule is described as the “number of non-water atoms observed experimentally in the asymmetric unit” ^[1] . Bucandin contains 516 atoms, which is considerably smaller than rubisco’s 17908 atoms. The last of the three major macromolecules is the residue count, which is explained as “number of polymetric residues in the asymmetric unit, based on the fully deposited sequence” ^[1]. Bucandin, again dwarfed by rubisco, has 63 amino acid residues in its structure as compared to rubisco’s 2,392 amino acids. All in all, Bucandin is a relatively small protein when sized up against other substantial proteins such as rubisco.

Function

When discussing a “presynaptic neurotoxin” such as Bucandin, this is referring to a toxin within the Malayan krait’s venom that attacks the neuromuscular junctions that allow us to innervate a muscle or muscle groups. This innervation begins when an action potential reaches this junction and causes the neurotransmitter to release from motor neurons, which in turn releases acetylcholine, which is a small molecule neurotransmitter. The binding of this acetylcholine to the receptor eventually leads to muscle contraction. When a snake bites, this release of acetylcholine is inhibited and therefore muscle contraction cannot occur. This eventually leads to paralysis, and if it spreads to the right regions (for example, the diaphragm which aids in breathing) death may occur.

Bucandin belongs to a class of proteins called , being named as such due to how its structure looks, resembling three fingers on a human hand. Three-finger toxins are typically only found in snake venom. These three-finger toxins contain four conserved that are rooted in a central core of three beta strand loops. These toxins are usually somewhere between 60 and 74 amino acid residues long. Bucandin being 63 amino acid residues falls well between these margins. Although they represent a wide variety of biological effects on the human body, three-finger toxins are almost always neurotoxins that act on the acetylcholine receptors. Apart from the three-finger toxins, the structure of Bucandin also includes two antiparallel beta sheets that have two strands and four strands, meaning that one of the beta-sheets has two strands that are bound together, forming one beta sheet, and the other beta sheet has four of said strands that form the other.

NMR Spectroscopy and Electron Density Map

Nuclear magnetic resonance spectroscopy, or NMR spectroscopy, is a way of observing the purity and structure of a protein through the use of a magnetic field. We are able to determine its structure based on how the magnetic resonance reflects off of each atom, giving us locations of these atoms. This will give us the basic structure of the compound as a whole, from which we can delve deeper into the structure to find out more specifics about it. The solution NMR structure for Bucandin shows us that it has two beta sheets as well as the generic three-finger toxin structure. One of the beta-sheets was a standard two stranded beta sheet, while the other is a four-stranded beta sheet. The four-stranded structure found in Bucandin is unusual for three-finger toxins, but the resonance for this four-stranded beta sheet was well represented by the NMR spectroscopy of Bucandin, telling us that, although unusual, is a part of the structure of Bucandin.

The amino acid tryptophan has an aromatic hydrophobic side chain. These side chains, Trp27 and Trp36 are facing towards the tip of the middle loop of the amino acid residues, giving the molecule as a whole a little bit of flexibility, showing us that Bucandin is not rigid.

The structure was also solved through crystallography. An electron density map is used to model X-ray structure determination. In X-ray structure determination, we shoot X-rays at a crystal structure of a protein and catch the refracted rays on a film. This allows us to create the pattern which in turn gives us the X-rays along with the intensity of the X-rays that gives us the final structure of the protein. The electron density map of Bucandin gives us some insight into the structure of this protein. It shows how the different amino acids form peptide bonds and when their structures are oriented in a certain direction. For example, by using the electron density map, we can see that there is an asparagine attached to glutamate at one point in the structure. According to ^[2], “the resulting electron density maps were of outstanding quality and allowed the automated tracing of 61 of 63 amino acid residues, including their side chains, and the placement of 48 solvent molecules”. This high-resolution structure allows us to obtain a greater understanding of the general structure and properties of Bucandin to identify specific characteristics of the protein.