Sandbox Reserved 1101: Difference between revisions

'''5IZ2''' is the '''N-terminal domain (NTD)''' of a spider protein called '''Major ampullate Spidroin 1A (MaSp1A)''', coming from the ''Nephila Clavipes'' species. This protein is a component of dragline silk produced in the major ampullate gland of spiders<ref name="Atkison"/>. The NTD domain of MaSp1A plays a major role in their combination during silk production <ref name="Atkison"/>. Indeed, thanks to the NTD dimerisation, two MaSps can be connected, leading to the formation of '''fibers''' with exceptional physical and biochemical qualities <ref name="José">José Roberto Aparecido dos Santos-Pinto, Helen Andrade Arcuri, Helga Priewalder, Heliana Clara Salles, Mario Sergio Palma and Gert Lubec, 2015. Structural Model for the Spider Silk Protein Spidroin‑1, Journal of Proteome research, 14, p.3859-3870.</ref>. It is of biotechnological interest to deeply understand the NTD dimerisation mechanism for the production of artificial spider silk, which can lead to innovative biomaterials.

The study of the ''N. Clavipes'' NTD permits to compare its structure with other species thus to provide new insights into the mechanism of NTD dimerization. Moreover, silks produced from different spider breeds vary in physical properties such as toughness and elasticity. In this way, studying diverse species would allow to '''optimize artificial silk''' for different applications.

The process of the dragline fiber formation is the connection of '''soluble MaSp''' proteins into '''insoluble fibers'''. Indeed, MaSps are firstly secreted and stored in soluble form in the tail of the major ampullate gland which is located in the spider’s abdomen. On demand, they pass through the narrow duct where they experience mechanical and chemical forces that convert them into fibers.  

[[Image:Image:Major ampullate gland.jpeg|300px|center|thumb| Schematic of Major ampullate gland of spiders. From Andersson et al. (2013). Morphology and composition of the spider major ampullate gland and dragline silk. Biomacromolecules ''14'', 2945-2952.]]

Actually, they deal with a pH dropping, an alteration of ion concentrations and oxidation conditions, which occur gradually along the duct. These changes promote the connection of MaSps extremities (i.e. homo-dimerisation of C- and N-terminal domains) to form fibers. Finally, through flow rate and mechanical forces experienced in the duct, the fibers will agglomerate to create the '''dragline silk'''.

[[Image:MaSp assembly.jpg|300px|center|thumb| Model of MaSps assembly according to the pH conditions. From Gaines et al. (2010). Spidroin N-terminal domain promotes a pH-dependent association of silk proteins during self-assembly. J. Biol. Chem. ''285'', 40745-40753.]]

The MaSps are between 250 to 350 kDa <ref name="Cadle"/>. They are divided into three parts : '''C-terminal domain (CTD)''', '''repeat domain (RD)''', and '''N-terminal domain (NTD)'''.  

*'''Repeat domain (RD)''':

The MaSp sequence corresponds to more than 90% of RD <ref>PMID:19221522</ref>. The RD is a long, flexible, highly repetitive central domain. It varies greatly between the types of silks, which makes it responsible for their different properties.

MaSp1 contains poly-alanine (A)n motifs at the end of a repeat, as well as GA and GGX motifs where X is often A, Y, L, or Q <ref name="Cadle"/>. The poly-alanine motifs, usually present at the end of a repeat, form [https://en.wikipedia.org/wiki/Beta_sheet_ β-sheets] in the duct due to mechanical forces. The β-sheets will then line up in parallel, leading to the aggregation of the fibers. The GGX motifs form an amorphous matrix that connects the crystalline regions <ref>PMID:15556872</ref>.

 

*'''C-terminal domain (CTD)''':

The CTD is a '''non-repetitive sequence''' of about 150 amino acids <ref name="Cadle"/>. The sequence identity, secondary structure and overall physical properties of CTD is highly conserved across spider species <ref name="Cadle"/>. Its structure forms a bundle of five parallel α-helices. A single '''cysteine residue''' in the middle of its sequence is highly conserved and is responsible for the CTD homo-dimerisation. In other words, it allows the covalent connection between two CTDs through disulfide bond linkage.

The CTD also plays a role in the change of MaSps solubility according to its localisation in the gland <ref name="Cadle"/>. Indeed, it presents a high number of charged and polar amino acids present in its sequence. In this way, when the pH is neutral in the ampullate, the hydrophobic residues are buried within the core and the hydrophilic residues are exposed. This permits to keep the MaSps soluble, preventing early fiber aggregation. On the contrary, when the CTDs are in the duct with lower pH, the acidic residues switch from a negative to a neutral charge. This leads to an increase of hydrophobic interactions that help with the formation of β-sheets and thus MaSps precipitation.

 

 

*'''N-terminal domain (NTD)''':

This domain is the most highly conserved domain. NTD dimerises in the duct upon conditions change, which connects the MaSps to form fibers.

The dragline silk represents the “toughest biopolymer on Earth” <ref>PMID:24119078</ref>. It also shows other beneficial properties including high tensile strength, elasticity  and biodegradability. That being, dragline fibers can have many uses in medical and industrial fields.

Synthetic silk proteins are commonly produced by recombinant gene expression and gene mimicry <ref>PMID:21999996</ref>. They can be spontaneously optimised by altering their form, size and composition. Indeed, DNA sections in silk protein sequence can be rearranged, added to or subtracted from to change the characteristics of the formed protein.

For instance, silk proteins can be processed into many different forms such as fibers, sponges, films, capsules and gels (<ref name="Cadle"/>. Their biodegradability can also be altered as required to increase or reduce their degradation time <ref name="Cadle"/>.

In this way, the uses for spider silk can give rise to a wide range of novel materials.

As far as the medical field is concerned, spider silk is naturally biocompatibility. This allows its use for applications like drug release materials, cell graft scaffolds, neuron regeneration and cartilage repair <ref name="Cadle"/>. Moreover, the spider silk can be recombinantly engineered to produce an antimicrobial property, certainly useful in this sector <ref>PMID:21458065</ref>.