Aciniform Spidroin, a Constituent of Egg Case Sacs and Wrapping Silk Fibers from the Black Widow Spider Latrodectus hesperus

Spiders produce high performance fibers with diverse mechanical properties and biological functions. Molecular and biochemical studies of spider egg case silk have revealed that the main constituent of the large diameter fiber contains the fibroin TuSp1. Here we demonstrate by SDS-PAGE and protein silver staining the presence of a distinct ∼300-kDa polypeptide that is found in solubilized egg case sacs. Combining matrix-assisted laser desorption ionization tandem time-of-flight mass spectrometry and reverse genetics, we have isolated a novel gene called AcSp1-like and demonstrate that its protein product is assembled into the small diameter fibers of egg case sacs and wrapping silks from the black widow spider, Latrodectus hesperus. BLAST searches of the NCBInr protein data base using the amino acid sequence of AcSp1-like revealed similarity to AcSp1, an inferred protein proposed to be a component of wrapping silk. However, the AcSp1-like protein was found to display more nonuniformity in its internal iterated repeat modules than the putative AcSp1 fibroin. Real time quantitative PCR analysis demonstrates that the AcSp1-like gene displays an aciniform gland-restricted pattern of expression. The amino acid composition of the fibroins extracted from the luminal contents of the aciniform glands was remarkably similar to the predicted amino acid composition of the AcSp1-like protein, which supports the assertion that AcSp1-like protein represents the major constituent stored within the aciniform gland. Collectively, our findings provide the first direct molecular evidence for the involvement of the aciniform gland in the production of a common fibroin that is assembled into the small diameter threads of egg case and wrapping silk of cob weavers. Spiders produce high performance fibers with diverse mechanical properties and biological functions. Molecular and biochemical studies of spider egg case silk have revealed that the main constituent of the large diameter fiber contains the fibroin TuSp1. Here we demonstrate by SDS-PAGE and protein silver staining the presence of a distinct ∼300-kDa polypeptide that is found in solubilized egg case sacs. Combining matrix-assisted laser desorption ionization tandem time-of-flight mass spectrometry and reverse genetics, we have isolated a novel gene called AcSp1-like and demonstrate that its protein product is assembled into the small diameter fibers of egg case sacs and wrapping silks from the black widow spider, Latrodectus hesperus. BLAST searches of the NCBInr protein data base using the amino acid sequence of AcSp1-like revealed similarity to AcSp1, an inferred protein proposed to be a component of wrapping silk. However, the AcSp1-like protein was found to display more nonuniformity in its internal iterated repeat modules than the putative AcSp1 fibroin. Real time quantitative PCR analysis demonstrates that the AcSp1-like gene displays an aciniform gland-restricted pattern of expression. The amino acid composition of the fibroins extracted from the luminal contents of the aciniform glands was remarkably similar to the predicted amino acid composition of the AcSp1-like protein, which supports the assertion that AcSp1-like protein represents the major constituent stored within the aciniform gland. Collectively, our findings provide the first direct molecular evidence for the involvement of the aciniform gland in the production of a common fibroin that is assembled into the small diameter threads of egg case and wrapping silk of cob weavers. The ability to spin multiple task-specific silks is a defining feature of the diverse order Araneae (>37,000 described species). Araneoid spiders use specialized abdominal glands to manufacture up to seven different protein-based silks/glues that have diverse mechanical properties (1Foelix R. Biology of Spider. Oxford University Press, New York1996: 1-336Google Scholar). Spinning high performance fibers with different mechanical properties enable spiders to perform a wide range of functions, including prey capture, locomotion, and protection of developing offspring (2Schultz J.W. Biol. Rev. 1987; 62: 89-113Crossref Google Scholar). Amino acid sequences of spider fibroins (spidroins) share a number of distinctive features. Repeats of four fundamental amino acid motifs characterize the majority of sequenced spider silks as follows: (i) alternating glycine and alanine ((GA)n), (ii) polyalanine (An), (iii) GGX (X = subset of residues), and (iv) GPGGX. Biochemical studies indicate that these motifs correspond to distinct structural modules, e.g. An and (GA)n repeats form crystalline β-sheets, whereas β-spirals are generated from a series of concatenated β-turns from the repeat structure GPGGX (3Hayashi C. Lewis R.V. J. Mol. Biol. 1998; 275: 773-784Crossref PubMed Scopus (339) Google Scholar). It has been proposed that these different structural modules contribute to the mechanics of the fibers. Combinations of these motifs form larger repetitive units termed ensemble repeats, which are organized in tandem copies throughout the silk sequence. In addition to ensemble repeat architecture, all spidroins have nonrepetitive C and N termini that exhibit length and sequence conservation (4Challis R.J. Goodacre S.L. Hewitt G.M. Insect Mol. Biol. 2006; 15: 45-56Crossref PubMed Scopus (51) Google Scholar, 5Sponner A. Unger E. Grosse F. Weisshart K. Biomacromolecules. 2004; 5: 840-845Crossref PubMed Scopus (68) Google Scholar, 6Motriuk-Smith D. Smith A. Hayashi C.Y. Lewis R.V. Biomacromolecules. 2005; 6: 3152-3159Crossref PubMed Scopus (101) Google Scholar, 7Hu X. Vasanthavada K. Kohler K. McNary S. Moore A.M. Vierra C.A. Cell. Mol. Life Sci. 2006; 63: 1986-1999Crossref PubMed Scopus (118) Google Scholar, 8Rising A. Hjalm G. Engstrom W. Johansson J. Biomacromolecules. 2006; 7: 3120-3124Crossref PubMed Scopus (125) Google Scholar). The C terminus has been postulated to play a role in the assembly process of spider silks (9Sponner A. Vater W. Rommerskirch W. Vollrath F. Unger E. Grosse F. Weisshart K. Biochem. Biophys. Res. Commun. 2005; 338: 897-902Crossref PubMed Scopus (72) Google Scholar, 10Huemmerich D. Helsen C.W. Quedzuweit S. Oschmann J. Rudolph R. Scheibel T. Biochemistry. 2004; 43: 13604-13612Crossref PubMed Scopus (306) Google Scholar). Spiders construct egg cases to serve as protective cocoons for their offspring (2Schultz J.W. Biol. Rev. 1987; 62: 89-113Crossref Google Scholar). Egg sacs must be designed to resist damage from predator/parasitoid invasion as well as temperature and humidity fluctuations (11Hieber C. J. Therm. Biol. 1985; 10: 171-175Crossref Scopus (34) Google Scholar, 12Hieber C. Oecologia. 1992; 91: 530-535Crossref PubMed Scopus (53) Google Scholar, 13Austin A.D. J. Nat. Hist. 1985; 19: 359-376Crossref Scopus (121) Google Scholar). Analyses of the physical structure of egg sacs collected from the cob weaver Latrodectus hesperus, commonly referred to as the black widow spider, have demonstrated that their egg case sacs consist of two different diameter fibers (14Hu X. Kohler K. Falick A.M. Moore A.M. Jones P.R. Sparkman O.D. Vierra C. J. Biol. Chem. 2005; 280: 21220-21230Abstract Full Text Full Text PDF PubMed Scopus (52) Google Scholar, 15Moon M.J. Korean J. Biol. Sci. 2003; 7: 35-41Crossref Google Scholar). MS/MS 4The abbreviations used are: MS/MS, tandem mass spectrometry; MALDI, matrix-assisted laser desorption ionization; TOF, time of flight; CAD, collision-activated dissociation; BLAST, basic local alignment search tool; AcSp1-like, aciniform spidroin 1-like; GdnHCl, guanidine hydrochloride; GdnSCN, guanidine thiocyanate; ORF, open reading frame; HPLC, high performance liquid chromatography. analyses of black widow peptides generated from solubilized egg case materials digested with trypsin have shown that the large diameter core fibers contain the fibroin TuSp1, as well as the egg case proteins ECP-1 and ECP-2 (egg case protein 1 or 2), which form a trimeric complex in cob weavers (14Hu X. Kohler K. Falick A.M. Moore A.M. Jones P.R. Sparkman O.D. Vierra C. J. Biol. Chem. 2005; 280: 21220-21230Abstract Full Text Full Text PDF PubMed Scopus (52) Google Scholar, 16Hu X. Kohler K. Falick A.M. Moore A.M. Jones P.R. Vierra C. Biochemistry. 2006; 45: 3506-3516Crossref PubMed Scopus (46) Google Scholar, 17Hu X. Lawrence B. Kohler K. Falick A.M. Moore A.M. McMullen E. Jones P.R. Vierra C. Biochemistry. 2005; 44: 10020-10027Crossref PubMed Scopus (56) Google Scholar). Real time quantitative PCR and Northern blot analyses have demonstrated tubuliform-restricted patterns of expression for these gene products (14Hu X. Kohler K. Falick A.M. Moore A.M. Jones P.R. Sparkman O.D. Vierra C. J. Biol. Chem. 2005; 280: 21220-21230Abstract Full Text Full Text PDF PubMed Scopus (52) Google Scholar, 17Hu X. Lawrence B. Kohler K. Falick A.M. Moore A.M. McMullen E. Jones P.R. Vierra C. Biochemistry. 2005; 44: 10020-10027Crossref PubMed Scopus (56) Google Scholar, 18Tian M. Lewis R.V. Biochemistry. 2005; 44: 8006-8012Crossref PubMed Scopus (63) Google Scholar, 19Zhao A.C. Zhao T.F. Nakagaki K. Zhang Y.S. Sima K. M. Nakagaki M. Biochemistry. 2006; 45: PubMed Scopus (56) Google Scholar). has been by the silk that the aciniform glands produce egg case silk and wrapping molecular evidence has been to demonstrate that fibroins are assembled into these diverse materials (1Foelix R. Biology of Spider. Oxford University Press, New York1996: 1-336Google Scholar). has been postulated that AcSp1, an inferred protein by the aciniform represents a main constituent of the luminal from aciniform evidence to the that fibroin is and assembled into wrapping fibers of weavers was C.Y. Lewis R.V. Mol. Biol. 2004; PubMed Scopus Google Scholar). the aciniform glands have been in the production of egg case silk and wrapping we that spiders spin common into egg case silk and wrapping Here we demonstrate by mass spectrometry and reverse genetics, the presence of the first aciniform gland-restricted AcSp1-like, that is assembled into egg case silk as well as wrapping findings provide into the molecular of aciniform silks and that black widow spiders use a common fibroin for egg case production and wrapping silk. of and the Egg trypsin was in acid a of 1 of the ∼300-kDa was a of for New Scholar). the ∼300-kDa in was from the the into with with and in a The and with with and and digested for with of extracted with two of of a acid with to the of the solubilized wrapping silk as of wrapping silk in of was by for to trypsin the was by the addition of of of trypsin and the was extracted and with a as described mass with a mass in MS/MS by in MS/MS sequences by of the high from the of wrapping silk by in in a and collected a that was into the mass An of in was with as from the and MS/MS from of the AcSp1-like sequence from the sequence was used to an was by an or the the and the reverse a of AcSp1-like from a from black widow silk glands to the amino in The a from which the major and and the The was sequenced as described B. M. J. M. R. J. Vierra C. Biophys. PubMed Scopus Google Scholar). a larger acid AcSp1-like, we the using with a acid using the AcSp1-like as a Real PCR as described J. R. M. J. G. Vierra C.A. Res. PubMed Scopus Google Scholar). Real time PCR was using an products with and using and used for the analysis of AcSp1-like the and reverse and used for the and reverse and silk and egg case silk to a of with in a with an a with an of of the to the in the temperature and which was and Amino of and silk fibers to amino acid analysis the of University as described D. K. J. Biol. PubMed Scopus (56) Google Scholar). silk and luminal collected from the aciniform gland with to the constituent amino collected by aciniform glands with by for using of Amino with and by by high performance liquid with of the ∼300-kDa and major structural proteins in the small diameter silk fibers of egg case we the threads with to the small diameter fibers and to the of the large diameter threads X. Kohler K. Falick A.M. Moore A.M. Jones P.R. Vierra C. Biochemistry. 2006; 45: 3506-3516Crossref PubMed Scopus (46) Google Scholar). SDS-PAGE analysis and silver staining of the solubilized egg case revealed the presence of with an molecular mass of patterns the was used to the egg case studies have demonstrated that two of the proteins egg case protein gene products (14Hu X. Kohler K. Falick A.M. Moore A.M. Jones P.R. Sparkman O.D. Vierra C. J. Biol. Chem. 2005; 280: 21220-21230Abstract Full Text Full Text PDF PubMed Scopus (52) Google Scholar, 16Hu X. Kohler K. Falick A.M. Moore A.M. Jones P.R. Vierra C. Biochemistry. 2006; 45: 3506-3516Crossref PubMed Scopus (46) Google Scholar). The ∼300-kDa protein the major constituent of the large diameter TuSp1, which is these and is from fibroins are high molecular was the ∼300-kDa protein was found to contain by analysis peptides sequenced using high CAD, with of and The product of the peptides and are shown in The N terminus of as well as the N and C termini of be a of sequence was The amino acid sequence of the was of the sequences using the BLAST revealed similarity to in the NCBInr protein data sequences by mass spectrometry sequences from the ∼300-kDa protein and tandem mass The N termini of as well as the N and C termini of be The the for predicted by the of the AcSp1-like has a mass that is to the mass by mass mass in a of a Egg the that the ∼300-kDa protein, we a from collected from The was by PCR using that to the sequences from MS/MS analysis The and the reverse designed from the sequence a to amino acid larger was by of the using the sequence as a for acid of the sequence a open reading the amino of a spider fibroin number protein a predicted molecular mass of and a of with sequences the In BLAST searches the predicted amino acid the to two putative fibroins AcSp1, number AcSp1, number sequence similarity and pattern of we have product AcSp1-like, which is from the aciniform spidroin AcSp1-like was to the A. and AcSp1 amino acid with of and for fibroin that share a nonrepetitive C AcSp1-like was found to be to AcSp1 from A. and AcSp1-like internal iterated modules that amino acid in length However, the internal modules found in the AcSp1 which amino acid modules, AcSp1-like ensemble repeats sequence alignment of the two ensemble repeats found within the amino acid sequence of AcSp1-like to be with In the spidroin An and in the AcSp1-like GGX modules of the peptides sequenced by MS/MS to of the AcSp1-like and The with which be sequenced its N or C terminus by MS/MS, was found to a mass from the AcSp1-like protein with trypsin The sequence of the which was used to the for the of the was found as a within our sequence. However, a of the predicted AcSp1-like amino acid sequence with trypsin the The to a mass that to within our AcSp1-like sequence indicate that in the N terminus of the AcSp1-like amino acid sequence. of AcSp1-like to the our mass data that AcSp1-like is within egg case silk we AcSp1-like was found within the aciniform gland. time quantitative PCR analysis to the expression of AcSp1-like in aciniform glands from the black widow spider AcSp1-like by the in the aciniform with in the major and and glands The expression of AcSp1-like in the aciniform gland was found to be than the gland or silk gland The AcSp1-like a of of the proposed for the aciniform gland is the production of wrapping silk for prey (1Foelix R. Biology of Spider. Oxford University Press, New York1996: 1-336Google Scholar). the physical structure of wrapping silk in hesperus, we wrapping silk collected from in silk threads using of wrapping silk revealed the presence of two different diameter fibers The larger fibers whereas fibers in diameter of the small diameter fibers in the these threads fibers the from of the aciniform the of the small diameter fibers is with their by the aciniform gland. small diameter fibers in wrapping silk of similar to the egg case small diameter the that the aciniform gland threads in wrapping silk and egg case sacs C and that the AcSp1-like protein was in wrapping we these threads with to of the solubilized wrapping silk with trypsin revealed the presence of peptides by analysis The of two of the and to from the AcSp1-like protein in egg case silk. The product of these two peptides and their sequences in found to be to of peptides found in the small diameter egg case silk fibers the mass which we sequenced by MS/MS was to the sequence of with the of an the C terminus of a sequences for and to found within the AcSp1 sequence 1 and we to sequence of the major peptides and we the for of the peptides using that the egg case AcSp1-like protein was within wrapping we the from the AcSp1-like sequence or within a mass range of a to the from wrapping silk with analysis of the products generated from wrapping as from the AcSp1-like protein, which and peptides sequenced by peptides found within the AcSp1-like sequence a sequence to two distinct of the AcSp1-like sequence and to be to found within the AcSp1-like with a The which was to have the sequence two and its was by MS/MS analysis to or of the AcSp1-like sequence that these two the It is to or by MS/MS have which sequence MS/MS analysis be sequenced of the MS/MS of the AcSp1-like sequence with trypsin a that an mass and as well as the that from the by MS/MS and Collectively, these studies demonstrate that of wrapping by MS/MS in four peptides to in the of egg case AcSp1-like protein and 1 and four peptides from wrapping silk trypsin and which sequenced by MS/MS, to found within the AcSp1-like sequence. these data that wrapping silk and egg case threads contain the AcSp1-like sequences by mass spectrometry MS/MS analyses of generated from the of solubilized wrapping silk from supports the presence of the egg case AcSp1-like indicate that be by the MS/MS The N terminus of be by the MS/MS However, of the predicted amino acid sequence of AcSp1-like generated a with an mass and sequence to mass AcSp1-like and contains a or peptides that found in the of the egg case AcSp1-like protein, as well as the of wrapping peptides that found in the of the egg case AcSp1-like protein, as well as the of wrapping contains a or and contains a or peptides that found in the of the egg case AcSp1-like protein, as well as the of wrapping contains a or peptides that found in the of the egg case AcSp1-like protein, as well as the of wrapping peptides that found in the of the egg case AcSp1-like protein, as well as the of wrapping contains a or peptides that found in the of the egg case AcSp1-like protein, as well as the of wrapping silk in a Amino of AcSp1-like, and spider silk glands are specialized designed to large of we the luminal contents of aciniform glands for the presence of the AcSp1-like The predicted amino acid composition of AcSp1-like was similar to the amino acid generated from the luminal collected from the aciniform gland that AcSp1-like represents of the major by the aciniform of the amino acid composition of the fibroins found in the wrapping from the amino acid composition of AcSp1-like of glycine and alanine found in wrapping silk to the stored aciniform fibroins In wrapping silk glycine and alanine and the presence of fiber The presence of fiber in wrapping silk was with the which revealed the presence of aciniform fibers as well as an larger diameter fiber amino acid composition of AcSp1-like and AcSp1, as well as the composition of wrapping silk egg case silk and egg case silk egg case of The are as an of multiple for egg case silk of are for for A. using the AcSp1 amino acid sequence number The are as and the and AcSp1-like of AcSp1 of A. are from the predicted amino acid sequence of A. AcSp1 using the and aciniform silk glands of = wrapping silk of = egg case silk of = are from egg case silk of = are from are from the predicted amino acid sequence of A. AcSp1 C.Y. Lewis R.V. Mol. Biol. 2004; PubMed Scopus Google using the E. C. A. S. A. The Google are from (14Hu X. Kohler K. Falick A.M. Moore A.M. Jones P.R. Sparkman O.D. Vierra C. J. Biol. Chem. 2005; 280: 21220-21230Abstract Full Text Full Text PDF PubMed Scopus (52) Google in a is by the spider silk that the aciniform glands to produce silks that are assembled into egg case and wrapping molecular evidence to assertion has been and reverse genetics, we have isolated a novel sequence that a of the fibroin that to the aciniform quantitative time PCR we demonstrate that AcSp1-like are in the aciniform gland than in MS/MS analyses of generated from egg case and wrapping silk fibers provide evidence for the presence of the AcSp1-like protein in assertion is by the of the peptides the sequence as the and from the of the egg case AcSp1-like protein and the of wrapping silk. the of from the black widow spider, we the that wrapping silk contains a AcSp1-like with large of amino acid sequence that are has been that the silk fibroin contain similar modules and that are assembled into larger ensemble repeats, their are of the amino acid sequences of the and fibroins with trypsin Hayashi C.Y. PubMed Scopus Google which are spidroins their expression similar amino acid and modules, produce with Combining the and molecular biological our findings provide the first evidence to the aciniform gland in the of a common fibroin that is in the small diameter fibers of egg case and wrapping silks of hesperus. these studies provide into the biochemical of aciniform that the AcSp1-like protein of the major that is and from the aciniform gland to produce that form a of egg case and wrapping BLAST searches of the NCBInr protein data base AcSp1-like protein revealed similarity to AcSp1, an inferred protein from the spider C.Y. Lewis R.V. Mol. Biol. 2004; PubMed Scopus Google Scholar). that the black widow AcSp1-like be as the to the inferred However, with the AcSp1, the AcSp1-like protein has distinctive including internal that are than in AcSp1 a acid which was found to be in the A. AcSp1 repeat modules C.Y. Lewis R.V. Mol. Biol. 2004; PubMed Scopus Google was from the amino acid sequence of The predicted amino acid composition of AcSp1-like and A. AcSp1, in including of and glycine in the AcSp1-like protein we are to the AcSp1-like gene as the AcSp1 in amino acid in the silk fibroins and which have similar predicted amino acid have been shown to be by distinct Hayashi C.Y. PubMed Scopus Google Scholar). BLAST searches with the AcSp1-like sequence the NCBInr data base similarity to the A. AcSp1 acid sequence. is in to the large diameter core fibroin TuSp1, which conservation the and amino acid for the and that our for AcSp1-like, we to that similar to AcSp1 gene is of the of conservation of sequence the the AcSp1 and AcSp1-like to the AcSp1-like gene has from the AcSp1 or AcSp1-like represents a distinct of the sequences from these silk fibers are of two distinct the and proteins in a within major silks A. Unger E. Grosse F. Weisshart K. Nat. 2005; PubMed Scopus Google is to that aciniform threads be from AcSp1 and AcSp1-like However, biochemical evidence to is with our of the predicted amino acid sequence of A. AcSp1 with trypsin to peptides that to found in digested black widow wrapping silk The to predicted A. in our solubilized wrapping silk digested with trypsin be by the the of AcSp1 in black widow spider fibers or the in the amino acid sequences of AcSp1 in and which the in our amino acid composition analysis of the aciniform silk proteins of A. that has of and than to the predicted amino acid composition of the inferred AcSp1 protein C.Y. Lewis R.V. Mol. Biol. 2004; PubMed Scopus Google Scholar). that the inferred AcSp1 protein, be stored with as the amino acid composition of the AcSp1-like protein to the fibroin amino acid composition of the luminal contents extracted from the aciniform glands of A. and hesperus. supports the assertion that fibroin is of the major stored in the aciniform the process of aciniform silks in hesperus. to be the AcSp1 in the aciniform glands of A. are into fibroins that are assembled into egg case and wrapping the of of and in the gland Hayashi C.Y. Sci. S. A. 2005; PubMed Scopus Google the to these fibroins in silks by MS/MS analyses of peptides generated from solubilized egg cases (14Hu X. Kohler K. Falick A.M. Moore A.M. Jones P.R. Sparkman O.D. Vierra C. J. Biol. Chem. 2005; 280: 21220-21230Abstract Full Text Full Text PDF PubMed Scopus (52) Google Scholar, 16Hu X. Kohler K. Falick A.M. Moore A.M. Jones P.R. Vierra C. Biochemistry. 2006; 45: 3506-3516Crossref PubMed Scopus (46) Google Scholar, 17Hu X. Lawrence B. Kohler K. Falick A.M. Moore A.M. McMullen E. Jones P.R. Vierra C. Biochemistry. 2005; 44: 10020-10027Crossref PubMed Scopus (56) Google the of fibroin are the of the spider that serve biological functions. to be the AcSp1 in weavers are and assembled into wrapping silk for prey capture, egg or silk studies of black widow egg cases with have shown that the large diameter fibers consist of a trimeric complex of TuSp1, and ECP-2 X. Kohler K. Falick A.M. Moore A.M. Jones P.R. Vierra C. Biochemistry. 2006; 45: 3506-3516Crossref PubMed Scopus (46) Google Scholar). Egg case fibers are with called and which have been in egg cases X. J. X. Vasanthavada K. Falick A.M. Jones P.R. C. Vierra C.A. Biochemistry. PubMed Scopus Google Scholar). The small diameter fibers of egg cases be solubilized by whereas the large diameter fibers are more to X. Kohler K. Falick A.M. Moore A.M. Jones P.R. Vierra C. Biochemistry. 2006; 45: 3506-3516Crossref PubMed Scopus (46) Google as well as of the the assertion that the AcSp1-like protein is assembled into the small diameter fibers of egg The AcSp1-like protein be in the solubilized of egg case which the small diameter fibers. The diameter of the aciniform to the diameter of the fibers. The AcSp1-like is in the aciniform gland. the presence of similar diameter fibers in wrapping silk is with the AcSp1-like protein to the small diameter fibers of wrapping the of the large diameter fibers in wrapping silk is However, the similarity of the diameter to that of major silks and the high glycine and alanine of wrapping is that these large diameter fibers are major with silk threads these fibers. of peptides generated by by sequence analysis using MS/MS, to in the solubilized wrapping silk are from peptides that are of the major silk fibroins or as aciniform silks these fibers are into egg cases and wrapping two different distinct biological functions. our we that aciniform silks serve to large diameter fibers which their presence in egg case and wrapping of the molecular mechanics and the of aciniform silks provide into the presence of aciniform silks in these the reading of the by of the of