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. 1989 Aug 1;109(2):607–618. doi: 10.1083/jcb.109.2.607

The Dictyostelium gelation factor shares a putative actin binding site with alpha-actinins and dystrophin and also has a rod domain containing six 100-residue motifs that appear to have a cross-beta conformation

PMCID: PMC2115711  PMID: 2668299

Abstract

The 120-kD gelation factor and alpha-actinin are among the most abundant F-actin cross-linking proteins in Dictyostelium discoideum. Both molecules are homodimers and have extended rod-like configurations that are respectively approximately 35 and 40 nm long. Here we report the complete cDNA sequence of the 120-kD gelation factor which codes for a protein of 857 amino acids. Its calculated molecular mass is 92.2 kD which is considerably smaller than suggested by its mobility in SDS- PAGE. Analysis of the sequence shows a region that is highly homologous to D. discoideum alpha-actinin, chicken fibroblast alpha-actinin, and human dystrophin. This conserved domain probably represents an actin binding site that is connected to the rod-forming part of the molecule via a highly charged stretch of amino acids. Whereas the sequence of alpha-actinin (Noegel, A., W. Witke, and M. Schleicher. 1987. FEBS [Fed. Eur. Biochem. Soc.] Lett. 221:391-396) suggests that the extended rod domain of the molecule is based on four spectrin-like repeats with high alpha-helix potential, the rod domain of the 120-kD gelation factor is constructed from six 100-residue repeats that have a high content of glycine and proline residues and which, in contrast to alpha- actinin, do not appear to have a high alpha-helical content. These repeats show a distinctive pattern of regions that have high beta-sheet potential alternating with short zones rich in residues with a high potential for turns. This observation suggests that each 100-residue motif has a cross-beta conformation with approximately nine sheets arranged perpendicular to the long axis of the molecule. In the high beta-potential zones every second residue is often hydrophobic. In a cross-beta structure, this pattern would result in one side of the domain having a surface rich in hydrophobic side chains which could account for the dimerization of the 120-kD gelation factor subunits.

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Selected References

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  1. André E., Lottspeich F., Schleicher M., Noegel A. Severin, gelsolin, and villin share a homologous sequence in regions presumed to contain F-actin severing domains. J Biol Chem. 1988 Jan 15;263(2):722–727. [PubMed] [Google Scholar]
  2. Baron M. D., Davison M. D., Jones P., Critchley D. R. The sequence of chick alpha-actinin reveals homologies to spectrin and calmodulin. J Biol Chem. 1987 Dec 25;262(36):17623–17629. [PubMed] [Google Scholar]
  3. Brier J., Fechheimer M., Swanson J., Taylor D. L. Abundance, relative gelation activity, and distribution of the 95,000-dalton actin-binding protein from Dictyostelium discoideum. J Cell Biol. 1983 Jul;97(1):178–185. doi: 10.1083/jcb.97.1.178. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Carboni J. M., Condeelis J. S. Ligand-induced changes in the location of actin, myosin, 95K (alpha-actinin), and 120K protein in amebae of Dictyostelium discoideum. J Cell Biol. 1985 Jun;100(6):1884–1893. doi: 10.1083/jcb.100.6.1884. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Chothia C. Coiling of beta-pleated sheets. J Mol Biol. 1983 Jan 5;163(1):107–117. doi: 10.1016/0022-2836(83)90031-1. [DOI] [PubMed] [Google Scholar]
  6. Chothia C., Janin J. Relative orientation of close-packed beta-pleated sheets in proteins. Proc Natl Acad Sci U S A. 1981 Jul;78(7):4146–4150. doi: 10.1073/pnas.78.7.4146. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Chou K. C., Némethy G., Rumsey S., Tuttle R. W., Scheraga H. A. Interactions between two beta-sheets. Energetics of beta/beta packing in proteins. J Mol Biol. 1986 Apr 20;188(4):641–649. doi: 10.1016/s0022-2836(86)80012-2. [DOI] [PubMed] [Google Scholar]
  8. Chou P. Y., Fasman G. D. Empirical predictions of protein conformation. Annu Rev Biochem. 1978;47:251–276. doi: 10.1146/annurev.bi.47.070178.001343. [DOI] [PubMed] [Google Scholar]
  9. Claviez M., Pagh K., Maruta H., Baltes W., Fisher P., Gerisch G. Electron microscopic mapping of monoclonal antibodies on the tail region of Dictyostelium myosin. EMBO J. 1982;1(8):1017–1022. doi: 10.1002/j.1460-2075.1982.tb01287.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Condeelis J., Vahey M. A calcium- and pH-regulated protein from Dictyostelium discoideum that cross-links actin filaments. J Cell Biol. 1982 Aug;94(2):466–471. doi: 10.1083/jcb.94.2.466. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Condeelis J., Vahey M., Carboni J. M., DeMey J., Ogihara S. Properties of the 120,000- and 95,000-dalton actin-binding proteins from Dictyostelium discoideum and their possible functions in assembling the cytoplasmic matrix. J Cell Biol. 1984 Jul;99(1 Pt 2):119s–126s. doi: 10.1083/jcb.99.1.119s. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Davison M. D., Critchley D. R. alpha-Actinins and the DMD protein contain spectrin-like repeats. Cell. 1988 Jan 29;52(2):159–160. doi: 10.1016/0092-8674(88)90503-x. [DOI] [PubMed] [Google Scholar]
  13. Emini E. A., Hughes J. V., Perlow D. S., Boger J. Induction of hepatitis A virus-neutralizing antibody by a virus-specific synthetic peptide. J Virol. 1985 Sep;55(3):836–839. doi: 10.1128/jvi.55.3.836-839.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Garnier J., Osguthorpe D. J., Robson B. Analysis of the accuracy and implications of simple methods for predicting the secondary structure of globular proteins. J Mol Biol. 1978 Mar 25;120(1):97–120. doi: 10.1016/0022-2836(78)90297-8. [DOI] [PubMed] [Google Scholar]
  15. Geddes A. J., Parker K. D., Atkins E. D., Beighton E. "Cross-beta" conformation in proteins. J Mol Biol. 1968 Mar 14;32(2):343–358. doi: 10.1016/0022-2836(68)90014-4. [DOI] [PubMed] [Google Scholar]
  16. Green N. M., Wrigley N. G., Russell W. C., Martin S. R., McLachlan A. D. Evidence for a repeating cross-beta sheet structure in the adenovirus fibre. EMBO J. 1983;2(8):1357–1365. doi: 10.1002/j.1460-2075.1983.tb01592.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Hammonds R. G., Jr Protein sequence of DMD gene is related to actin-binding domain of alpha-actinin. Cell. 1987 Oct 9;51(1):1–1. doi: 10.1016/0092-8674(87)90002-x. [DOI] [PubMed] [Google Scholar]
  18. Hamodrakas S. J., Bosshard H. E., Carlson C. N. Structural models of the evolutionarily conservative central domain of silk-moth chorion proteins. Protein Eng. 1988 Sep;2(3):201–207. doi: 10.1093/protein/2.3.201. [DOI] [PubMed] [Google Scholar]
  19. Henikoff S. Unidirectional digestion with exonuclease III creates targeted breakpoints for DNA sequencing. Gene. 1984 Jun;28(3):351–359. doi: 10.1016/0378-1119(84)90153-7. [DOI] [PubMed] [Google Scholar]
  20. Imamura M., Endo T., Kuroda M., Tanaka T., Masaki T. Substructure and higher structure of chicken smooth muscle alpha-actinin molecule. J Biol Chem. 1988 Jun 5;263(16):7800–7805. [PubMed] [Google Scholar]
  21. Kaufmann E., Geisler N., Weber K. SDS-PAGE strongly overestimates the molecular masses of the neurofilament proteins. FEBS Lett. 1984 May 7;170(1):81–84. doi: 10.1016/0014-5793(84)81373-3. [DOI] [PubMed] [Google Scholar]
  22. Koenig M., Monaco A. P., Kunkel L. M. The complete sequence of dystrophin predicts a rod-shaped cytoskeletal protein. Cell. 1988 Apr 22;53(2):219–228. doi: 10.1016/0092-8674(88)90383-2. [DOI] [PubMed] [Google Scholar]
  23. Lacombe M. L., Podgorski G. J., Franke J., Kessin R. H. Molecular cloning and developmental expression of the cyclic nucleotide phosphodiesterase gene of Dictyostelium discoideum. J Biol Chem. 1986 Dec 25;261(36):16811–16817. [PubMed] [Google Scholar]
  24. Lee G., Cowan N., Kirschner M. The primary structure and heterogeneity of tau protein from mouse brain. Science. 1988 Jan 15;239(4837):285–288. doi: 10.1126/science.3122323. [DOI] [PubMed] [Google Scholar]
  25. Lipman D. J., Pearson W. R. Rapid and sensitive protein similarity searches. Science. 1985 Mar 22;227(4693):1435–1441. doi: 10.1126/science.2983426. [DOI] [PubMed] [Google Scholar]
  26. MacLean-Fletcher S. D., Pollard T. D. Viscometric analysis of the gelation of Acanthamoeba extracts and purification of two gelation factors. J Cell Biol. 1980 May;85(2):414–428. doi: 10.1083/jcb.85.2.414. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Maizel J. V., Jr, Lenk R. P. Enhanced graphic matrix analysis of nucleic acid and protein sequences. Proc Natl Acad Sci U S A. 1981 Dec;78(12):7665–7669. doi: 10.1073/pnas.78.12.7665. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. McLachlan A. D., Stewart M. The 14-fold periodicity in alpha-tropomyosin and the interaction with actin. J Mol Biol. 1976 May 15;103(2):271–298. doi: 10.1016/0022-2836(76)90313-2. [DOI] [PubMed] [Google Scholar]
  29. Newell P. C., Telser A., Sussman M. Alternative developmental pathways determined by environmental conditions in the cellular slime mold Dictyostelium discoideum. J Bacteriol. 1969 Nov;100(2):763–768. doi: 10.1128/jb.100.2.763-768.1969. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Noegel A. A., Witke W. Inactivation of the alpha-actinin gene in Dictyostelium. Dev Genet. 1988;9(4-5):531–538. doi: 10.1002/dvg.1020090429. [DOI] [PubMed] [Google Scholar]
  31. Noegel A., Welker D. L., Metz B. A., Williams K. L. Presence of nuclear associated plasmids in the lower eukaryote Dictyostelium discoideum. J Mol Biol. 1985 Sep 20;185(2):447–450. doi: 10.1016/0022-2836(85)90416-4. [DOI] [PubMed] [Google Scholar]
  32. Noegel A., Witke W., Schleicher M. Calcium-sensitive non-muscle alpha-actinin contains EF-hand structures and highly conserved regions. FEBS Lett. 1987 Sep 14;221(2):391–396. doi: 10.1016/0014-5793(87)80962-6. [DOI] [PubMed] [Google Scholar]
  33. Pollard T. D., Cooper J. A. Actin and actin-binding proteins. A critical evaluation of mechanisms and functions. Annu Rev Biochem. 1986;55:987–1035. doi: 10.1146/annurev.bi.55.070186.005011. [DOI] [PubMed] [Google Scholar]
  34. Pollard T. D., Thomas S. M., Niederman R. Human platelet myosin. I. Purification by a rapid method applicable to other nonmuscle cells. Anal Biochem. 1974 Jul;60(1):258–266. doi: 10.1016/0003-2697(74)90152-3. [DOI] [PubMed] [Google Scholar]
  35. Salemme F. R. Structural properties of protein beta-sheets. Prog Biophys Mol Biol. 1983;42(2-3):95–133. doi: 10.1016/0079-6107(83)90005-6. [DOI] [PubMed] [Google Scholar]
  36. Salisbury J. L., Condeelis J. S., Maihle N. J., Satir P. Calmodulin localization during capping and receptor-mediated endocytosis. Nature. 1981 Nov 12;294(5837):163–166. doi: 10.1038/294163a0. [DOI] [PubMed] [Google Scholar]
  37. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Scheel J., Ziegelbauer K., Kupke T., Humbel B. M., Noegel A. A., Gerisch G., Schleicher M. Hisactophilin, a histidine-rich actin-binding protein from Dictyostelium discoideum. J Biol Chem. 1989 Feb 15;264(5):2832–2839. [PubMed] [Google Scholar]
  39. Schleicher M., André E., Hartmann H., Noegel A. A. Actin-binding proteins are conserved from slime molds to man. Dev Genet. 1988;9(4-5):521–530. doi: 10.1002/dvg.1020090428. [DOI] [PubMed] [Google Scholar]
  40. Schleicher M., Gerisch G., Isenberg G. New actin-binding proteins from Dictyostelium discoideum. EMBO J. 1984 Sep;3(9):2095–2100. doi: 10.1002/j.1460-2075.1984.tb02096.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Schleicher M., Noegel A., Schwarz T., Wallraff E., Brink M., Faix J., Gerisch G., Isenberg G. A Dictyostelium mutant with severe defects in alpha-actinin: its characterization using cDNA probes and monoclonal antibodies. J Cell Sci. 1988 May;90(Pt 1):59–71. doi: 10.1242/jcs.90.1.59. [DOI] [PubMed] [Google Scholar]
  42. Simon M. N., Mutzel R., Mutzel H., Véron M. Vectors for expression of truncated coding sequences in Escherichia coli. Plasmid. 1988 Mar;19(2):94–102. doi: 10.1016/0147-619x(88)90048-0. [DOI] [PubMed] [Google Scholar]
  43. Sobue K., Kanda K., Tanaka T., Ueki N. Caldesmon: a common actin-linked regulatory protein in the smooth muscle and nonmuscle contractile system. J Cell Biochem. 1988 Jul;37(3):317–325. doi: 10.1002/jcb.240370306. [DOI] [PubMed] [Google Scholar]
  44. Speicher D. W., Marchesi V. T. Erythrocyte spectrin is comprised of many homologous triple helical segments. Nature. 1984 Sep 13;311(5982):177–180. doi: 10.1038/311177a0. [DOI] [PubMed] [Google Scholar]
  45. Speicher D. W. The present status of erythrocyte spectrin structure: the 106-residue repetitive structure is a basic feature of an entire class of proteins. J Cell Biochem. 1986;30(3):245–258. doi: 10.1002/jcb.240300306. [DOI] [PubMed] [Google Scholar]
  46. Spudich J. A., Watt S. The regulation of rabbit skeletal muscle contraction. I. Biochemical studies of the interaction of the tropomyosin-troponin complex with actin and the proteolytic fragments of myosin. J Biol Chem. 1971 Aug 10;246(15):4866–4871. [PubMed] [Google Scholar]
  47. Stewart M., Beveridge T. J., Sprott G. D. Crystalline order to high resolution in the sheath of Methanospirillum hungatei: a cross-beta structure. J Mol Biol. 1985 Jun 5;183(3):509–515. doi: 10.1016/0022-2836(85)90019-1. [DOI] [PubMed] [Google Scholar]
  48. Stewart M. Introduction to the computer image processing of electron micrographs of two-dimensionally ordered biological structures. J Electron Microsc Tech. 1988 Aug;9(4):301–324. doi: 10.1002/jemt.1060090403. [DOI] [PubMed] [Google Scholar]
  49. Stewart M., McLachlan A. D. Fourteen actin-binding sites on tropomyosin? Nature. 1975 Sep 25;257(5524):331–333. doi: 10.1038/257331a0. [DOI] [PubMed] [Google Scholar]
  50. Stewart M. The structure of chicken scale keratin. J Ultrastruct Res. 1977 Jul;60(1):27–33. doi: 10.1016/s0022-5320(77)80038-5. [DOI] [PubMed] [Google Scholar]
  51. Stossel T. P., Chaponnier C., Ezzell R. M., Hartwig J. H., Janmey P. A., Kwiatkowski D. J., Lind S. E., Smith D. B., Southwick F. S., Yin H. L. Nonmuscle actin-binding proteins. Annu Rev Cell Biol. 1985;1:353–402. doi: 10.1146/annurev.cb.01.110185.002033. [DOI] [PubMed] [Google Scholar]
  52. Towbin H., Staehelin T., Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350–4354. doi: 10.1073/pnas.76.9.4350. [DOI] [PMC free article] [PubMed] [Google Scholar]
  53. Wallraff E., Schleicher M., Modersitzki M., Rieger D., Isenberg G., Gerisch G. Selection of Dictyostelium mutants defective in cytoskeletal proteins: use of an antibody that binds to the ends of alpha-actinin rods. EMBO J. 1986 Jan;5(1):61–67. doi: 10.1002/j.1460-2075.1986.tb04178.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  54. Warrick H. M., Spudich J. A. Codon preference in Dictyostelium discoideum. Nucleic Acids Res. 1988 Jul 25;16(14A):6617–6635. doi: 10.1093/nar/16.14.6617. [DOI] [PMC free article] [PubMed] [Google Scholar]
  55. Wasenius V. M., Närvänen O., Lehto V. P., Saraste M. Alpha-actinin and spectrin have common structural domains. FEBS Lett. 1987 Aug 31;221(1):73–76. doi: 10.1016/0014-5793(87)80354-x. [DOI] [PubMed] [Google Scholar]
  56. Witke W., Nellen W., Noegel A. Homologous recombination in the Dictyostelium alpha-actinin gene leads to an altered mRNA and lack of the protein. EMBO J. 1987 Dec 20;6(13):4143–4148. doi: 10.1002/j.1460-2075.1987.tb02760.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  57. Witke W., Schleicher M., Lottspeich F., Noegel A. Studies on the transcription, translation, and structure of alpha-actinin in Dictyostelium discoideum. J Cell Biol. 1986 Sep;103(3):969–975. doi: 10.1083/jcb.103.3.969. [DOI] [PMC free article] [PubMed] [Google Scholar]
  58. Yanisch-Perron C., Vieira J., Messing J. Improved M13 phage cloning vectors and host strains: nucleotide sequences of the M13mp18 and pUC19 vectors. Gene. 1985;33(1):103–119. doi: 10.1016/0378-1119(85)90120-9. [DOI] [PubMed] [Google Scholar]
  59. Young R. A., Davis R. W. Efficient isolation of genes by using antibody probes. Proc Natl Acad Sci U S A. 1983 Mar;80(5):1194–1198. doi: 10.1073/pnas.80.5.1194. [DOI] [PMC free article] [PubMed] [Google Scholar]

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