Conservation of Essential Design Features in Coiled Coil Silks

Abstract

Silks are strong protein fibers produced by a broad array of spiders and insects. The vast majority of known silks are large, repetitive proteins assembled into extended beta-sheet structures. Honeybees, however, have found a radically different evolutionary solution to the need for a building material. The 4 fibrous proteins of honeybee silk are small ( approximately 30 kDa each) and nonrepetitive and adopt a coiled coil structure. We examined silks from the 3 superfamilies of the Aculeata (Hymenoptera: Apocrita) by infrared spectroscopy and found coiled coil structure in bees (Apoidea) and in ants (Vespoidea) but not in parasitic wasps of the Chrysidoidea. We subsequently identified and sequenced the silk genes of bumblebees, bulldog ants, and weaver ants and compared these with honeybee silk genes. Each species produced orthologues of the 4 small fibroin proteins identified in honeybee silk. Each fibroin contained a continuous predicted coiled coil region of around 210 residues, flanked by 23-160 residue length N- and C-termini. The cores of the coiled coils were unusually rich in alanine. There was extensive sequence divergence among the bee and ant silk genes (<50% similarity between the alignable regions of bee and ant sequences), consistent with constant and equivalent divergence since the bee/ant split (estimated to be 155 Myr). Despite a high background level of sequence diversity, we have identified conserved design elements that we propose are essential to the assembly and function of coiled coil silks.

Physical properties of honeybee silk: a review

“…Sutherland et al (2006) were able to identify the coiled-coil silk sequences from silk gland cDNA libraries from European A. mellifera and determined the amino acid sequence of the coiled-coils. Sutherland et al (2007) confirmed that honeybee silk is formed from four coiled-coil proteins (fibroins), as originally suggested by Rudall (1962Rudall ( , 1965 on the basis of his X-ray diffraction data. The fibroin proteins contained extensive coiled-coil regions of conserved length flanked by largely unstructured termini.…”

“…The patterns from honeybee silk fibres were considered most consistent with a four-strand coiled-coil structure with a tighter than expected super-helix radius of about 0.52 nm (Atkins 1967). In contrast, the dominant molecular structure in silk of other hymenopteran species is extended β-sheets (Warwicker 1960;Sutherland et al 2007) So, honeybee silk is an ά-helical fibroin (Rudall 1962), the micelles or crystallites of which form a four-stranded array of coiled-coils parallel to the fibre axis (Atkins 1967). Honeybee fibroin is crystalline relative to other insect silks (Lucas and Rudall 1968) while the hydrated fibre is only half as stiff as dry ones although they are equal in strength (Hepburn et al 1979).…”

“…So, it is not surprising that the discovery of the amino acid sequence in honeybee silk protein provided an explanation of why the coiled-coil packing was atypically tight: While the core of coiled-coils usually contains large hydrophobic residues such as leucine and isoleucine, in coiled-coil silk the most abundant core residue is the small amino acid alanine (Sutherland et al 2007). Lucas and Rudall (1968) suggested that the pattern of coiled-coil proteins occurring in the silk gland could be to prevent agglutination of the proteins within the silk gland.…”

“…The fibroin proteins contained extensive coiled-coil regions of conserved length flanked by largely unstructured termini. Sutherland et al (2007) proposed a structural model for coiledcoil silks (Figure 5), whereby the ά-helical strands corresponding to each of the fibroins are arranged in an antiparallel tetrameric configuration (direction indicated by arrows).…”

“…Most known coiled-coils contain predominantly large hydrophobic residues at these positions to maximise the hydrophobic forces stabilising the core (Woolfson 2005). Sutherland et al (2007) ascribed the atypical composition of the coiled-coils in bee silks possibly to the metabolic constraints of having to produce a continuous and copious secretion of silk during the many hours of larval spinning.…”