Recombinant Spider Silk: Promises and Bottlenecks

Abstract

Spider silk threads have exceptional mechanical properties such as toughness, elasticity and low density, which reach maximum values compared to other fibre materials. They are superior even compared to Kevlar and steel. These extraordinary properties stem from long length and specific protein structures. Spider silk proteins can consist of more than 20,000 amino acids. Polypeptide stretches account for more than 90% of the whole protein, and these domains can be repeated more than a hundred times. Each repeat unit has a specific function resulting in the final properties of the silk. These properties make them attractive for innovative material development for medical or technical products as well as cosmetics. However, with livestock breeding of spiders it is not possible to reach high volumes of silk due to the cannibalistic behaviour of these animals. In order to obtain spider silk proteins (spidroins) on a large scale, recombinant production is attempted in various expression systems such as plants, bacteria, yeasts, insects, silkworms, mammalian cells and animals. For viable large-scale production, cost-effective and efficient production systems are needed. This review describes the different types of spider silk, their proteins and structures and discusses the production of these difficult-to-express proteins in different host organisms with an emphasis on plant systems.

Engineering a minimal SilkCatcher/Tag pair compatible with SpyCatcher/Tag pair for the production of native-sized spider silk

“…Large-scale production of spider silk proteins has been hindered by difficulties in the recombinant production of native-sized spider silk proteins in high yields and in a soluble form [21] . Attempts to express long dragline silk sequences in production organisms have generally resulted in truncated proteins, low expression yields, and/or insoluble proteins although the fibers spun from them have had promising properties [24,40,41] .…”

“…A widely interesting application of protein conjugation is multiple fragment ligation [15,18,19] that allows the recombinant production of long and repetitive proteins that are difficult to produce in the common production hosts, such as Escherichia coli. Spider silk is an important and interesting example of this, because of its extreme properties both at molecular and macroscopic level, as well as the numerous applications [20][21][22] . The desirable mechanical properties of spider-silk fibers, together with the unfeasibility of farming spiders, have prompted research aiming at producing recombinant spider silk.…”

“…Spider silk proteins, called spidroins, consist of very long (thousands of amino acids, 250-320 kDa) and highly repetitive alanine-and glycine-rich regions [23,24] . The size and repetitive nature of the core parts are crucial for the fiber properties but create significant challenges for the recombinant production of the spidroins [21,22] . Multiple fragment ligation is, however, limited by the lack of efficient and orthogonal (non-cross-reactive) protein ligation methods.…”

Molecular atlas reveals the tri-sectional spinning mechanism of spider dragline silk

“…Much of our current understanding of spider silk formation is based on physical and material studies that have provided only a partial picture of its nature 42, 43 . A comprehensive molecular biological atlas of the Ma gland and dragline silk is fundamental to uncovering how spiders make dragline silk fibers 11, 44 . Additionally, such a detailed basis of silk formation will be valuable for ultratough fiber innovation 5, 8, 42, 45 .…”

Recombinant protein condensation inside E. coli enables the development of building blocks for bioinspired materials engineering – biomimetic spider silk protein as a case study

“…In their native forms, these proteins often contain intrinsically disordered regions, long and repetitive sequences and/or amyloidogenic motifs. When such native sequences are recombinantly overexpressed, in most cases, they exhibit very low production yield or are prone to premature aggregation and accumulation into insoluble inclusion bodies [15,16,17,18]. Protein sequestration into inclusion bodies during recombinant expression in bacteria is a widely used strategy to achieve very high production yield [19,20].…”