A Novel Highly Divergent Protein Family Identified from a Viviparous Insect by RNA-seq Analysis: A Potential Target for Tsetse Fly-Specific Abortifacients

English version České info

Tsetse flies are the sole vector for African trypanosomes, causative agents of sleeping sickness in humans and nagana in cattle. Transcriptome and proteome analyses were utilized to examine the underlying mechanisms of tsetse lactation that occur during each reproductive cycle. These analyses revealed a dramatic shift to the synthesis of milk proteins during lactation and a novel milk-specific protein family. All members of this family were co-localized, shared sequence similarity and were expressed at 40× basal levels during milk secretion. Suppression of gene from this lactation-associated family impaired progeny development by reducing milk protein content and altering milk homeostasis. These novel genes represent an excellent target for tsetse-specific reproductive-based control mechanisms. In addition, the characterization of tsetse milk production revealed multiple factors that are functionally analogous between tsetse and mammalian lactation.

Vyšlo v časopise: A Novel Highly Divergent Protein Family Identified from a Viviparous Insect by RNA-seq Analysis: A Potential Target for Tsetse Fly-Specific Abortifacients. PLoS Genet 10(4): e32767. doi:10.1371/journal.pgen.1003874
Kategorie: Research Article
prolekare.web.journal.doi_sk: https://doi.org/10.1371/journal.pgen.1003874

Souhrn

Zdroje

1. TobeSS, LangleyPA (1978) Reproductive physiology of Glossina. Ann Rev Entomol 23 : 283–307.

2. CmelikSHW, BursellE, SlackE (1969) Composition of the gut contents of thrid-instar tsetse larvae (Glossina morsitans Westwood). Comp Biochem Physiol 29 : 447–453.

3. EjezieGC, DaveyKG (1976) Some effects of allatectomy in female tsetse, Glossina austeni. J Insect Physiol 22 : 1743–1749.

4. DenlingerDL (1975) Insect hormones as tsetse abortifacients. Nature 253 : 347–348.

5. DenlingerDL, MaW-C (1974) Dynamics of the pregnancy cycle in the tsetse Glossina morsitans. J Insect Physiol 20 : 1015–1026.

6. AttardoGM, GuzN, Strickler-DinglasanP, AksoyS (2006) Molecular aspects of viviparous reproductive biology of the tsetse fly (Glossina morsitans morsitans): Regulation of yolk and milk gland protein synthesis. J Insect Physiol 52 : 1128–1136.

7. GuzN, AttardoGM, WuY, AksoyS (2007) Molecular aspects of transferrin expression in the tsetse fly (Glossina morsitans morsitans). J Insect Physiol 53 : 715–723.

8. AttardoGM, RibeiroJMC, WuYN, BerrimanM, AksoyS (2010) Transcriptome analysis of reproductive tissue and intrauterine developmental stages of the tsetse fly (Glossina morsitans morsitans). BMC Genomics 11: .

9. BenoitJB, AttardoGM, MichalkovaV, TakacP, BohovaJ, et al. (2012) Sphingomyelinase activity in mother's milk is essential for juvenile development: a case from lactating tsetse flies. Biol Reprod 87 : 1–10.

10. Strickler-DinglasanPM, GuzN, AttardoG, AksoyS (2006) Molecular characterization of iron binding proteins from Glossina morsitans morsitans (Diptera : Glossinidae). Insect Biochem Mol Biol 36 : 921–933.

11. OsirEO, KotengoM, ChaudhuryMFB, OtienoLH (1991) Structural studies on the major milk gland protein of the tsetse fly, Glossina morsitans morsitans. Comp Biochem Physiol B-Biochem 99 : 803–809.

12. YangG, AttardoGM, LohsC, AksoyS (2010) Molecular characterization of two novel milk proteins in the tsetse fly (Glossina morsitans morsitans). Insect Mol Biol 19 : 253–262.

13. WangJ, AksoyS (2012) PGRP-LB is a maternally transmitted immune milk protein that influences symbiosis and parasitism in tsetse's offspring. Proc Natl Acad Sci USA 109 : 10552–10557.

14. BaumannA, BenoitJB, MichalkovaV, MirejiPO, AttardoGM, et al. (2012) Interactions between juvenile hormone and insulin signaling pathways regulate lipid levels during lactation and dry periods associated with tsetse fly pregnancy. Mol Cell Endocrinol 372 : 30–41.

15. International Glossina Genome Initiative (2013) Genome sequence of the tsetse fly (Glossina morsitans): vector of African trypanosomiasis. Science In preparation.

16. BirolI, JackmanSD, NielsenCB, QianJQ, VarholR, et al. (2009) De novo transcriptome assembly with ABySS. Bioinformatics 25 : 2872–2877.

17. RobertsonG, ScheinJ, ChiuR, CorbettR, FieldM, et al. (2010) De novo assembly and analysis of RNA-seq data. Nat Methods 7 : 909–912.

18. GrabherrMG, HaasBJ, YassourM, LevinJZ, ThompsonDA, et al. (2011) Full-length transcriptome assembly from RNA-Seq data without a reference genome. Nat Biotechnol 29 : 644–652.

19. TsygankovAY (2009) TULA-family proteins: an odd couple. Cell Mol Life Sci 66 : 2949–2952.

20. RaglandGJ, DenlingerDL, HahnDA (2010) Mechanisms of suspended animation are revealed by transcript profiling of diapause in the flesh fly. Proc Natl Acad Sci U S A 107 : 14909–14914.

21. HahnDA, RaglandGJ, ShoemakerDD, DenlingerDL (2009) Gene discovery using massively parallel pyrosequencing to develop ESTs for the flesh fly Sarcophaga crassipalpis. BMC Genomics 10 : 234.

22. IkaiA (1980) Thermostability and aliphatic index of globular proteins. J Biochem 88 : 1895–1898.

23. ThompsonJD, GibsonTJ, PlewniakF, JeanmouginF, HiggginsDG (1997) The CLUSTAL_X windows interface: flexible strategies for multiple sequence alignment aided by quality analysis tools. Nuc Acids Res 25 : 4876–4882.

24. HallTA (1999) BioEdit: a user-friendly biological sequence alignment editor and analysis program for Windows 95/98/NT. Nuc Acids Sym Ser 41 : 95–98.

25. TamuraK, DudleyJ, NeiM, KumarS (2007) MEGA4: Molecular Evolutionary Genetics Analysis (MEGA) software version 4.0. Mol Biol Evol 24 : 1596–1599.

26. TamuraK, PetersonD, PetersonN, StecherG, NeiM, et al. (2011) MEGA5: molecular evolutionary genetics analysis using maximum likelihood, evolutionary distance, and maximum parsimony methods. Mol Biol Evol 28 : 2731–2739.

27. SchefflerK, MartinDP, SeoigheC (2006) Robust inference of positive selection from recombining coding sequences. Bioinformatics 22 : 2493–2499.

28. PondSLK, FrostSDW (2005) Not so different after all: A comparison of methods for detecting amino acid sites under selection. Mol Biol Evol 22 : 1208–1222.

29. DelportW, PoonAFY, FrostSDW, PondSLK (2010) Datamonkey 2010: a suite of phylogenetic analysis tools for evolutionary biology. Bioinformatics 26 : 2455–2457.

30. PondSLK, FrostSDW (2005) Datamonkey: rapid detection of selective pressure on individual sites of codon alignments. Bioinformatics 21 : 2531–2533.

31. PondSLK, FrostSDW, MuseSV (2005) HyPhy: hypothesis testing using phylogenies. Bioinformatics 21 : 676–679.

32. PharoEA, De LeoAA, RenfreeMB, ThomsonPC, LefevreCM, et al. (2012) The mammary gland-specific marsupial ELP and eutherian CTI share a common ancestral gene. BMC Evol Biol 12 : 80.

33. MaWC, DenlingerDL, JarlforsU, SmithDS (1975) Structural modulations in the tsetse fly milk gland during a pregnancy cycle. Tissue & Cell 7 : 319–330.

34. Bonnanfant-JaisML (1974) Morphologie de la gland lactee d-une glossine, Glossina austeni Newst, au cours du cycle de gestation. I. Aspects ultrastructuraux en periode de gestation J Microscopy 19 : 265–284.

35. LenobleBJ, DenlingerDL (1982) The milk gland of the sheep ked, Melophagus ovinus -⁠ a comparison with Glossina. J Insect Physiol 28 : 165–167.

36. TobeSS, DaveyKG (1974) Autoradiographic study of protein synthesis in abdominal tissues of Glossina austeni. Tissue & Cell 6 : 255–268.

37. LangleyPA, BursellE (1980) Role of fat body and uterine gland in milk synthesis by adult female Glossina morsitans. Insect Biochem 10 : 11–17.

38. RiddifordLM, DhadiallaTS (1990) Protein synthesis by the milk gland and fat body of the tsetse fly, Glossina pallidipes. Insect Biochem 20 : 493–500.

39. LefevreCM, DigbyMR, WhitleyJC, StrahmY, NicholasKR (2007) Lactation transcriptomics in the Australian marsupial, Macropus eugenii: transcript sequencing and quantification. BMC Genomics 8 : 417.

40. MarinottiO, CalvoE, NguyenQK, DissanayakeS, RibeiroJM, et al. (2006) Genome-wide analysis of gene expression in adult Anopheles gambiae. Insect Mol Biol 15 : 1–12.

41. ParisiM, NuttallR, EdwardsP, MinorJ, NaimanD, et al. (2004) A survey of ovary-, testis-, and soma-biased gene expression in Drosophila melanogaster adults. Genome Biol 5: R40.

42. McGrawLA, ClarkAG, WolfnerMF (2008) Post-mating gene expression profiles of female Drosophila melanogaster in response to time and to four male accessory gland proteins. Genetics 179 : 1395–1408.

43. BionazM, PeriasamyK, Rodriguez-ZasSL, EvertsRE, LewinHA, et al. (2012) Old and new stories: revelations from functional analysis of the bovine mammary transcriptome during the lactation cycle. PLoS One 7: e33268.

44. LemayDG, NevilleMC, RudolphMC, PollardKS, GermanJB (2007) Gene regulatory networks in lactation: identification of global principles using bioinformatics. BMC Syst Biol 1 : 56.

45. PatelOV, CaseyT, DoverH, PlautK (2011) Homeorhetic adaptation to lactation: comparative transcriptome analysis of mammary, liver, and adipose tissue during the transition from pregnancy to lactation in rats. Funct Integr Genomics 11 : 193–202.

46. WickramasingheS, RinconG, Islas-TrejoA, MedranoJF (2012) Transcriptional profiling of bovine milk using RNA sequencing. BMC Genomics 13 : 45.

47. AttardoGM, BenoitJB, MichalkovaV, YangG, RollerL, et al. (2012) Analysis of lipolysis underlying lactation in the tsetse fly, Glossina morsitans. Insect Biochem Mol Biol 42 : 360–370.

48. WangB, MoyaN, NiessenS, HooverH, MihaylovaMM, et al. (2011) A hormone-dependent module regulating energy balance. Cell 145 : 596–606.

49. BellerM, BulankinaAV, HsiaoHH, UrlaubH, JackleH, et al. (2010) PERILIPIN-dependent control of lipid droplet structure and fat storage in Drosophila. Cell Metabolism 12 : 521–532.

50. BellerM, ThielK, ThulPJ, JackleH (2010) Lipid droplets: A dynamic organelle moves into focus. FEBS Letters 584 : 2176–2182.

51. TelemanAA (2010) Molecular mechanisms of metabolic regulation by insulin in Drosophila. Biochem J 425 : 13–26.

52. EllisJE, ParkerL, ChoJ, AroraK (2010) Activin signaling functions upstream of Gbb to regulate synaptic growth at the Drosophila neuromuscular junction. Dev Biol 342 : 121–133.

53. ClarkRI, WoodcockKJ, GeissmannF, TrouilletC, DionneMS (2011) Multiple TGF-beta superfamily signals modulate the adult Drosophila immune response. Curr Biol 21 : 1672–1677.

54. BloiseE, CassaliGD, FerreiraMC, CiarmelaP, PetragliaF, et al. (2010) Activin-related proteins in bovine mammary gland: localization and differential expression during gestational development and differentiation. J Dairy Sci 93 : 4592–4601.

55. CiarmelaP, BloiseE, GrayPC, CarrarelliP, IslamMS, et al. (2011) Activin-A and myostatin response and steroid regulation in human myometrium: disruption of their signalling in uterine fibroid. J Clin Endocrinol Metab 96 : 755–765.

56. RobinsonGW, HennighausenL (1997) Inhibins and activins regulate mammary epithelial cell differentiation through mesenchymal-epithelial interactions. Development 124 : 2701–2708.

57. BursellE (1977) Synthesis of proline by the fat body of the tsetse fly (Glossina morsitans). metabolic pathways. Insect Biochem 7 : 427–434.

58. AttardoGM, Strickler-DinglasanP, PerkinSAH, CalerE, BonaldoMF, et al. (2006) Analysis of fat body transcriptome from the adult tsetse fly, Glossina morsitans morsitans. Insect Mol Biol 15 : 411–424.

59. PimleyRW, LangleyPA (1982) Hormone stimulated lipolysis and proline synthesis in the fat body of the adult tsetse fly, Glossina morsitans. J Insect Physiol 28 : 781–789.

60. KrahmerN, GuoY, WilflingF, HilgerM, LingrellS, et al. (2011) Phosphatidylcholine synthesis for lipid droplet expansion is mediated by localized activation of CTP:phosphocholine cytidylyltransferase. Cell Metabolism 14 : 504–515.

61. BlusztajnJK (1998) Choline, a vital amine. Science 281 : 794–795.

62. VanderzantES (1974) Development, significance, and application of artificial diets for insects. Ann Rev Entomol 19 : 139–160.

63. RigdenDJ (2008) The histidine phosphatase superfamily: structure and function. Biochem J 409 : 333–348.

64. TsygankovAY (2013) TULA-family proteins: a new class of cellular regulators. J Cell Physiol 228 : 43–49.

65. DenlingerDL, MaWC (1975) Maternal nutritive secretions as possible channels for vertical transmission of microorganisms in insects: the tsetse fly example. Ann NY Acad Sci 266 : 162–165.

66. AttardoGM, LohsC, HeddiA, AlamUH, YildirimS, et al. (2008) Analysis of milk gland structure and function in Glossina morsitans: milk protein production, symbiont populations and fecundity. J Insect Physiol 54 : 1236–1242.

67. WeissBL, MaltzM, AksoyS (2012) Obligate symbionts activate immune system development in the tsetse fly. J Immunol 188 : 3395–3403.

68. WeissBL, WangJ, MaltzMA, AksoyS (2013) Trypanosome infection establishment in the tsetse is influenced by microbiome-regulated host immune barrier. PLoS Pathogens 9: e1003318.

69. RijnkelsM (2002) Multispecies comparison of the casein gene loci and evolution of casein gene family. J Mammary Gland Biol Neoplasia 7 : 327–345.

70. GingerMR, GrigorMR (1999) Comparative aspects of milk caseins. Comp Biochem Physiol B 124 : 133–145.

71. CampbellEM, BallA, HopplerS, BowmanAS (2008) Invertebrate aquaporins: a review. J Comp Physiol B 178 : 935–955.

72. ZardoyaR (2005) Phylogeny and evolution of the major intrinsic protein family. Biol Cell 97 : 397–414.

73. GalindoK, SmithDP (2001) A large family of divergent Drosophila odorant-binding proteins expressed in gustatory and olfactory sensilla. Genetics 159 : 1059–1072.

74. GrahamLA, DaviesPL (2002) The odorant-binding proteins of Drosophila melanogaster: annotation and characterization of a divergent gene family. Gene 292 : 43–55.

75. CaspersGJ, LeunissenJA, de JongWW (1995) The expanding small heat-shock protein family, and structure predictions of the conserved “alpha-crystallin domain”. J Mol Evol 40 : 238–248.

76. McManamanJL, NevilleMC (2003) Mammary physiology and milk secretion. Adv Drug Del Rev 55 : 629–641.

77. NevilleMC, PiccianoMF (1997) Regulation of milk lipid secretion and composition. Ann Rev Nut 17 : 159–183.

78. LemayDG, LynnDJ, MartinWF, NevilleMC, CaseyTM, et al. (2009) The bovine lactation genome: insights into the evolution of mammalian milk. Genome Biol 10: R43.

79. O'DonnellR, HollandJW, DeethHC, AlewoodP (2004) Milk proteomics. Int Dairy J 14 : 1013–1023.

80. KontopidisG, HoltC, SawyerL (2004) Invited review: beta-lactoglobulin: binding properties, structure, and function. J Dairy Sci 87 : 785–796.

81. NybergL, FarooqiA, BlackbergL, DuanRD, NilssonA, et al. (1998) Digestion of ceramide by human milk bile salt-stimulated lipase. J Pediatr Gastroenterol Nutr 27 : 560–567.

82. DuanRD (2011) Physiological functions and clinical implications of sphingolipids in the gut. J Digest Dis 12 : 60–70.

83. DuanRD (2007) Sphingomyelinase and ceramidase in the intestinal tract. Eur J Lipid Sci Tech 109 : 987–993.

84. ClarksonRW, WaylandMT, LeeJ, FreemanT, WatsonCJ (2004) Gene expression profiling of mammary gland development reveals putative roles for death receptors and immune mediators in post-lactational regression. Breast Cancer Res 6: R92–109.

85. HettingaK, van ValenbergH, de VriesS, BoerenS, van HooijdonkT, et al. (2011) The host defense proteome of human and bovine milk. PLoS One 6: e19433.

86. DenlingerDL, MaWC (1975) Maternal nutritive secretions as possible channels for vertical transmission of microorganisms in insects: the tsetse fly example. Ann N Y Acad Sci 266 : 162–165.

87. Lara-VillosladaF, OlivaresM, SierraS, RodriguezJM, BozaJ, et al. (2007) Beneficial effects of probiotic bacteria isolated from breast milk. Br J Nutr 98 Suppl 1: S96–100.

88. MartinR, LangaS, ReviriegoC, JiminezE, MarinML, et al. (2003) Human milk is a source of lactic acid bacteria for the infant gut. J Pediatr 143 : 754–758.

89. MangeA, BelletV, TuaillonE, de PerrePV, SolassolJ (2008) Comprehensive proteomic analysis of the human milk proteome: contribution of protein fractionation. J Chromat B-Anal Tech Biomed Life Sci 876 : 252–256.

90. ZivkovicAM, GermanJB, LebrillaCB, MillsDA (2011) Human milk glycobiome and its impact on the infant gastrointestinal microbiota. Proc Natl Acad Sci U S A 108 Suppl 1 : 4653–4658.

91. HollandJW, DeethHC, AlewoodPF (2004) Proteomic analysis of kappa-casein micro-heterogeneity. Proteomics 4 : 743–752.

92. LefevreCM, SharpJA, NicholasKR (2010) Evolution of lactation: ancient origin and extreme adaptations of the lactation system. Annu Rev Genomics Hum Genet 11 : 219–238.

93. DickinsonE (2006) Structure formation in casein-based gels, foams, and emulsions Colloids and Surfaces A: Physicochem. Eng Aspects 288 : 3–11.

94. ShekarPC, GoelS, RaniSDS, SarathiDP, AlexJL, et al. (2006) kappa-Casein-deficient mice fail to lactate. Proc Natl Acad Sci U S A 103 : 8000–8005.

95. KumarS, ClarkeAR, HooperML, HorneDS, LawAJR, et al. (1994) Milk composition and lactation of Beta casein deficient mice. Proc Natl Acad Sci U S A 91 : 11767–11767.

96. KolbAF, HuberRC, LillicoSG, CarlisleA, RobinsonCJ, et al. (2011) Milk lacking alpha-casein leads to permanent reduction in body size in mice. PLoS One 6.

97. ChanatE, MartinP, Ollivier-BousquetM (1999) alpha (S1)-casein is required for the efficient transport of beta -⁠ and kappa-casein from the endoplasmic reticulum to the Golgi apparatus of mammary epithelial cells. J Cell Sci 112 : 3399–3412.

98. RousseauD (2000) Fat crystals and emulsion stability -⁠ a review. Food Res Int 33 : 3–14.

99. AllenJC, WriedenWL (1982) Influence of milk proteins on lipid oxidation in aqueous emulsion .1. casein, whey protein and alpha lactalbumin. J Dairy Res 49 : 239–248.

100. FongBY, NorrisCS, MacGibbonAKH (2007) Protein and lipid composition of bovine milk-fat-globule membrane. Int Dairy J 17 : 275–288.

101. VanderghemC, BleckerC, DanthineS, DeroanneC, HaubrugeE, et al. (2008) Proteome analysis of the bovine milk fat globule: enhancement of membrane purification. Int Dairy J 18 : 885–893.

102. CharlwoodJ, HanrahanS, TyldesleyR, LangridgeJ, DwekM, et al. (2002) Use of proteomic methodology for the characterization of human milk fat globular membrane proteins. Anal Biochem 301 : 314–324.

103. CeboC, LopezC, HenryC, BeauvalletC, MenardO, et al. (2012) Goat alpha(s1)-casein genotype affects milk fat globule physicochemical properties and the composition of the milk fat globule membrane. J Dairy Sci 95 : 6215–6229.

104. MolooSK (1971) An artificial feeding technique for Glossina. Parasitology 63 : 507–512.

105. DenlingerDL (1972) Induction and termination of pupal diapause in Sarcophaga (Diptera: Sarcophagidae). Biol Bull 142 : 11–24.

106. RioRV, SymulaRE, WangJ, LohsC, WuYN, et al. (2012) Insight into the transmission biology and species-specific functional capabilities of tsetse (Diptera: Glossinidae) obligate symbiont Wigglesworthia. mBio 3.

107. TohH, WeissBL, PerkinSA, YamashitaA, OshimaK, et al. (2006) Massive genome erosion and functional adaptations provide insights into the symbiotic lifestyle of Sodalis glossinidius in the tsetse host. Genome Res 16 : 149–156.

108. AltschulSF, MaddenTL, SchafferAA, ZhangJ, ZhangZ, et al. (1997) Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res 25 : 3389–3402.

109. TatusovRL, FedorovaND, JacksonJD, JacobsAR, KiryutinB, et al. (2003) The COG database: an updated version includes eukaryotes. BMC Bioinformatics 4 : 41.

110. AshburnerM, BallCA, BlakeJA, BotsteinD, ButlerH, et al. (2000) Gene ontology: tool for the unification of biology. The Gene Ontology Consortium. Nat Genet 25 : 25–29.

111. SchafferAA, AravindL, MaddenTL, ShavirinS, SpougeJL, et al. (2001) Improving the accuracy of PSI-BLAST protein database searches with composition-based statistics and other refinements. Nucleic Acids Res 29 : 2994–3005.

112. BatemanA, BirneyE, DurbinR, EddySR, HoweKL, et al. (2000) The Pfam protein families database. Nucleic Acids Res 28 : 263–266.

113. NielsenH, EngelbrechtJ, BrunakS, von HeijneG (1997) Identification of prokaryotic and eukaryotic signal peptides and prediction of their cleavage sites. Protein Eng 10 : 1–6.

114. TweedieS, AshburnerM, FallsK, LeylandP, McQuiltonP, et al. (2009) FlyBase: enhancing Drosophila Gene Ontology annotations. Nucleic Acids Res 37: D555–559.

115. LawsonD, ArensburgerP, AtkinsonP, BesanskyNJ, BruggnerRV, et al. (2007) VectorBase: a home for invertebrate vectors of human pathogens. Nucleic Acids Res 35: D503–505.

116. MortazaviA, WilliamsBA, McCueK, SchaefferL, WoldB (2008) Mapping and quantifying mammalian transcriptomes by RNA-Seq. Nat Methods 5 : 621–628.

117. KalAJ, van ZonneveldAJ, BenesV, van den BergM, KoerkampMG, et al. (1999) Dynamics of gene expression revealed by comparison of serial analysis of gene expression transcript profiles from yeast grown on two different carbon sources. Mol Biol Cell 10 : 1859–1872.

118. HirosawaM, HoshidaM, IshikawaM, ToyaT (1993) MASCOT: multiple alignment system for protein sequences based on three-way dynamic programming. Comput Appl Biosci 9 : 161–167.

119. PappinDJ, HojrupP, BleasbyAJ (1993) Rapid identification of proteins by peptide-mass fingerprinting. Curr Biol 3 : 327–332.

120. PerkinsDN, PappinDJ, CreasyDM, CottrellJS (1999) Probability-based protein identification by searching sequence databases using mass spectrometry data. Electrophoresis 20 : 3551–3567.

121. IshihamaY, OdaY, TabataT, SatoT, NagasuT, et al. (2005) Exponentially modified protein abundance index (emPAI) for estimation of absolute protein amount in proteomics by the number of sequenced peptides per protein. Mol Cell Proteomics 4 : 1265–1272.

122. ShinodaK, TomitaM, IshihamaY (2010) emPAI Calc–for the estimation of protein abundance from large-scale identification data by liquid chromatography-tandem mass spectrometry. Bioinformatics 26 : 576–577.

123. PeiJ, KimBH, GrishinNV (2008) PROMALS3D: a tool for multiple protein sequence and structure alignments. Nucleic Acids Res 36 : 2295–2300.

124. LarkinMA, BlackshieldsG, BrownNP, ChennaR, McGettiganPA, et al. (2007) Clustal W and Clustal X version 2.0. Bioinformatics 23 : 2947–2948.

125. KumarS, TamuraK, NeiM (2004) MEGA 3: Integrated software for molecular evolutionary genetics analysis and sequence alignment. Brief Bioinform 5 : 150–163.

126. XuD, ZhangY (2012) Ab initio protein structure assembly using continuous structure fragments and optimized knowledge-based force field. Proteins 80 : 1715–1735.

127. RoyA, KucukuralA, ZhangY (2010) I-TASSER: a unified platform for automated protein structure and function prediction. Nat Protoc 5 : 725–738.

128. KelleyLA, SternbergMJ (2009) Protein structure prediction on the Web: a case study using the Phyre server. Nat Protoc 4 : 363–371.

129. YangY, FaraggiE, ZhaoH, ZhouY (2011) Improving protein fold recognition and template-based modeling by employing probabilistic-based matching between predicted one-dimensional structural properties of query and corresponding native properties of templates. Bioinformatics 27 : 2076–2082.

130. BlomN, Sicheritz-PontenT, GuptaR, GammeltoftS, BrunakS (2004) Prediction of post-translational glycosylation and phosphorylation of proteins from the amino acid sequence. Proteomics 4 : 1633–1649.

131. JuleniusK, MolgaardA, GuptaR, BrunakS (2005) Prediction, conservation analysis, and structural characterization of mammalian mucin-type O-glycosylation sites. Glycobiology 15 : 153–164.

132. BlomN, GammeltoftS, BrunakS (1999) Sequence and structure-based prediction of eukaryotic protein phosphorylation sites. J Mol Biol 294 : 1351–1362.