Spider Minor Ampullate Silk Proteins Are Constituents of Prey Wrapping Silk in the Cob WeaverLatrodectus hesperus

Abstract

Spiders spin high performance fibers with diverse biological functions and mechanical properties. Molecular and biochemical studies of spider prey wrapping silks have revealed the presence of the aciniform silk fibroin AcSp1-like. In our studies we demonstrate the presence of a second distinct polypeptide present within prey wrapping silk. Combining matrix-assisted laser desorption ionization tandem time-of-flight mass spectrometry and reverse genetics, we have isolated a novel gene called MiSp1-like and demonstrate that its protein product is a constituent of prey wrap silks from the black widow spider, Latrodectus hesperus. BLAST searches of the NCBInr protein database using the amino acid sequence of MiSp1-like revealed similarity to the conserved C-terminal domain of silk family members. In particular, MiSp1-like showed the highest degree of sequence similarity to the nonrepetitive C-termini of published orb-weaver minor ampullate fibroin molecules. Analysis of the internal amino acid sequence of the black widow MiSp1-like revealed polyalanine stretches interrupted by glycine residues and glycine-alanine couplets within MiSp1-like as well as repeats of the heptameric sequence AGGYGQG. Real-time quantitative PCR analysis demonstrates that the MiSp1-like gene displays a minor ampullate gland-restricted pattern of expression. Furthermore, amino acid composition analysis, coupled with scanning electron microscopy of raw wrapping silk, supports the assertion that minor ampullate silks are important constituents of black widow spider prey wrap silk. Collectively, our findings provide direct molecular evidence for the involvement of minor ampullate fibroins in swathing silks and suggest composite materials play an important role in the wrap attack process for cob-weavers.

Structure of silk by raman spectromicroscopy: From the spinning glands to the fibers

“…The silk used to wrap preys caught in the web is produced by the aciniform (Ac) glands, although it has been shown that, in addition to Ac fibers, Ma and Mi silk fibers are also found in the wrapping net spun around a prey. 56 Interestingly, Raman spectromicroscopy has clearly confirmed the above finding, showing that this technique is also an efficient and simple tool to identify the nature of silk fibers in harvested biological samples. 57 Until recently, no data was available regarding the structural organization of the Ac silk, but its amino acid sequence has been determined for Argiope trifasciata.…”

Pyriform Spidroin 1, a Novel Member of the Silk Gene Family That Anchors Dragline Silk Fibers in Attachment Discs of the Black Widow Spider, Latrodectus hesperus

“…A, major ampullate; B, minor ampullate; C, flagelliform; D, aggregate; E, aciniform; F, tubuliform; and G, pyriform. Data from the major and minor ampullate glands, as well as the aciniform gland were previously determined (6,12). Fig.…”

“…Over the past 20 years, seven distinct members of the silk gene family have been identified and characterized at the molecular level, which include the silk proteins MaSp1 and MaSp2 (dragline silk) (3)(4)(5), AcSp1 or AcSp1-like (wrapping silk and egg case silk) (6,7), TuSp1 (egg case silk)(8 -11), MiSp1 and MiSp2 or MiSp1-like (temporary spiral capture silk or web reinforcement silk) (12,13), and Flag silk (capture spiral silk in orb weavers) (14). These spider fibroins have revealed that they share a number of distinctive properties, including four fundamental amino acid repeat motifs that characterize the majority of the family: (i) alternating glycine alanine couplets (GA n ), (ii) polyalanine blocks (A n ), (iii) GGX (X ϭ subset of residues, which include Leu, Ile, and Ala), and (iv) GPGX n .…”

Solid-state nuclear magnetic resonance (NMR) spectroscopy reveals distinctive protein dynamics in closely related spider silks

“…34 It has also been found in the wrapping net spun around prey. 35 Cocoon silk is produced by the tubuliform gland of female spiders during the reproductive season 36 and is used to make the cocoon outer egg sac. As shown in Table 1, MiS has a lower stress at rupture than MaS, but its extensibility is higher.…”