De Novo Design, Expression, and Characterization of Felix: a Four-Helix Bundle Protein of Native-Like Sequence
10.1126/science.2392678
Crossref journal-article
American Association for the Advancement of Science (AAAS)
Science (221)
Abstract

The protein Felix was designed de novo to fold into an antiparallel four-helix bundle of specific topology. Its sequence of 79 amino acid residues is not homologous to any known protein sequence, but is "native-like" in that it is nonrepetitive and contains 19 of the 20 naturally occurring amino acids. Felix has been expressed from a synthetic gene cloned in Escherichia coli , and the protein has been purified to homogeneity. Physical characterization of the purified protein indicates that Felix (i) is monomeric in solution, (ii) is predominantly α-helical, (iii) contains a designed intramolecular disulfide bond linking the first and fourth helices, and (iv) buries its single tryptophan in an apolar environment and probably in close proximity with the disulfide bond. These physical properties rule out several alternative structures and indicate that Felix indeed folds into approximately the designed three-dimensional structure.

Bibliography

Hecht, M. H., Richardson, J. S., Richardson, D. C., & Ogden, R. C. (1990). De Novo Design, Expression, and Characterization of Felix: a Four-Helix Bundle Protein of Native-Like Sequence. Science, 249(4971), 884–891.

Authors 4
  1. Michael H. Hecht (first)
  2. Jane S. Richardson (additional)
  3. David C. Richardson (additional)
  4. Richard C. Ogden (additional)
References 65 Referenced 347
  1. 10.1073/pnas.84.18.6434
  2. ARGOS, P, 4-HELICAL SUPER-SECONDARY STRUCTURE, BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS 75: 83 (1977). (10.1016/0006-291X(77)91292-X) / BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS (1977)
  3. BANNER, D, JOURNAL OF MOLECULAR BIOLOGY 196: 657 (1987). (10.1016/0022-2836(87)90039-8) / JOURNAL OF MOLECULAR BIOLOGY (1987)
  4. BERNSTEIN, F.C., PROTEIN DATA BANK - COMPUTER-BASED ARCHIVAL FILE FOR MACROMOLECULAR STRUCTURES, JOURNAL OF MOLECULAR BIOLOGY 112: 535 (1977). (10.1016/S0022-2836(77)80200-3) / JOURNAL OF MOLECULAR BIOLOGY (1977)
  5. BRACK, A, MULTICONFORMATIONAL SYNTHETIC POLYPEPTIDES, JOURNAL OF THE AMERICAN CHEMICAL SOCIETY 103: 6319 (1981). (10.1021/ja00411a009) / JOURNAL OF THE AMERICAN CHEMICAL SOCIETY (1981)
  6. BRAHMS, S, DETERMINATION OF PROTEIN SECONDARY STRUCTURE IN SOLUTION BY VACUUM ULTRAVIOLET CIRCULAR-DICHROISM, JOURNAL OF MOLECULAR BIOLOGY 138: 149 (1980). (10.1016/0022-2836(80)90282-X) / JOURNAL OF MOLECULAR BIOLOGY (1980)
  7. 10.1126/science.3500515
  8. BULLOCK, P.A., CIRCULAR-DICHROISM AND RESONANCE RAMAN STUDIES OF CYTOCHROME-B562 FROM ESCHERICHIA-COLI, BIOCHEMISTRY 17: 3084 (1978). (10.1021/bi00608a022) / BIOCHEMISTRY (1978)
  9. CHOU, P.Y., Prediction of the Secondary Structure of Proteins from their Amino Acid Sequence, ADVANCES IN ENZYMOLOGY 47: 45 (1978). / ADVANCES IN ENZYMOLOGY / Prediction of the Secondary Structure of Proteins from their Amino Acid Sequence (1978)
  10. COWGILL, R.W., FLUORESCENCE AND STRUCTURE OF PROTEINS .18. SPATIAL REQUIREMENTS FOR QUENCHING BY DISULFIDE GROUPS, BIOCHIMICA ET BIOPHYSICA ACTA 207: 556 (1970). (10.1016/S0005-2795(70)80019-8) / BIOCHIMICA ET BIOPHYSICA ACTA (1970)
  11. DANIELS, S.B., CHEMICAL SYNTHESIS OF BETA-STRUCTURES - BETABELLIN-9, FASEB JOURNAL 2: A1543 (1988). / FASEB JOURNAL (1988)
  12. de Boer, H. A., Maximizing Gene Expression: 225 (1986). / Maximizing Gene Expression (1986)
  13. DIJKSTRA, B.W., ROLE OF THE N-TERMINUS IN THE INTERACTION OF PANCREATIC PHOSPHOLIPASE-A2 WITH AGGREGATED SUBSTRATES - PROPERTIES AND CRYSTAL-STRUCTURE OF TRANSAMINATED PHOSPHOLIPASE-A2, BIOCHEMISTRY 23: 2759 (1984). (10.1021/bi00307a035) / BIOCHEMISTRY (1984)
  14. EFIMOV, A.V., MOL BIOL 16: 271 (1982). / MOL BIOL (1982)
  15. EISENBERG, D, THE HELICAL HYDROPHOBIC MOMENT - A MEASURE OF THE AMPHIPHILICITY OF A HELIX, NATURE 299: 371 (1982). (10.1038/299371a0) / NATURE (1982)
  16. Eisenberg, D., Proceedings of the National Academy of Sciences of the United States of America 81: 140 (1984). / Proceedings of the National Academy of Sciences of the United States of America (1984)
  17. EISENBERG, D, PROTEIN-STRUCT FUNCT 1: 16 (1986). (10.1002/prot.340010105) / PROTEIN-STRUCT FUNCT (1986)
  18. FINZEL, B.C., STRUCTURE OF FERRICYTOCHROME C' FROM RHODOSPIRILLUM-MOLISCHIANUM AT 1.67-A RESOLUTION, JOURNAL OF MOLECULAR BIOLOGY 186: 627 (1985). (10.1016/0022-2836(85)90135-4) / JOURNAL OF MOLECULAR BIOLOGY (1985)
  19. GARNIER, J, ANALYSIS OF ACCURACY AND IMPLICATIONS OF SIMPLE METHODS FOR PREDICTING SECONDARY STRUCTURE OF GLOBULAR PROTEINS, JOURNAL OF MOLECULAR BIOLOGY 120: 97 (1978). (10.1016/0022-2836(78)90297-8) / JOURNAL OF MOLECULAR BIOLOGY (1978)
  20. GREENFIELD, N, COMPUTED CIRCULAR DICHROISM SPECTRA FOR EVALUATION OF PROTEIN CONFORMATION, BIOCHEMISTRY 8: 4108 (1969). (10.1021/bi00838a031) / BIOCHEMISTRY (1969)
  21. HECHT, M.H., J CELL BIOCHEM 13: 86 (1989). / J CELL BIOCHEM (1989)
  22. HENDRICKSON, W.A., PSEUDO-SYMMETRY IN STRUCTURE OF MYOHEMERYTHRIN, JOURNAL OF BIOLOGICAL CHEMISTRY 252: 3012 (1977). (10.1016/S0021-9258(17)40465-0) / JOURNAL OF BIOLOGICAL CHEMISTRY (1977)
  23. HENDRICKSON, W.A., STRUCTURE OF THE HYDROPHOBIC PROTEIN CRAMBIN DETERMINED DIRECTLY FROM THE ANOMALOUS SCATTERING OF SULFUR, NATURE 290: 107 (1981). (10.1038/290107a0) / NATURE (1981)
  24. HO, S.P., DESIGN OF A 4-HELIX BUNDLE PROTEIN - SYNTHESIS OF PEPTIDES WHICH SELF-ASSOCIATE INTO A HELICAL PROTEIN, JOURNAL OF THE AMERICAN CHEMICAL SOCIETY 109: 6751 (1987). (10.1021/ja00256a032) / JOURNAL OF THE AMERICAN CHEMICAL SOCIETY (1987)
  25. HOLLADAY, L.A., GROWTH-HORMONE CONFORMATION AND CONFORMATIONAL EQUILIBRIA, BIOCHEMISTRY 13: 1653 (1974). (10.1021/bi00705a015) / BIOCHEMISTRY (1974)
  26. HOLLEY, L.H., PROTEIN SECONDARY STRUCTURE PREDICTION WITH A NEURAL NETWORK, PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA 86: 152 (1989). / PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA (1989)
  27. ISENMAN, D.E., FOLDING PATHWAYS OF IMMUNOGLOBULIN DOMAINS - FOLDING KINETICS OF THE C-GAMMA-3 DOMAIN OF HUMAN IGGL, BIOCHEMISTRY 18: 3327 (1979). (10.1021/bi00582a020) / BIOCHEMISTRY (1979)
  28. JOHNSON, W.C., SECONDARY STRUCTURE OF PROTEINS THROUGH CIRCULAR-DICHROISM SPECTROSCOPY, ANNUAL REVIEW OF BIOPHYSICS AND BIOPHYSICAL CHEMISTRY 17: 145 (1988). (10.1146/annurev.bb.17.060188.001045) / ANNUAL REVIEW OF BIOPHYSICS AND BIOPHYSICAL CHEMISTRY (1988)
  29. KUWAJIMA, K, THE MOLTEN GLOBULE STATE AS A CLUE FOR UNDERSTANDING THE FOLDING AND COOPERATIVITY OF GLOBULAR-PROTEIN STRUCTURE, PROTEINS-STRUCTURE FUNCTION AND GENETICS 6: 87 (1989). (10.1002/prot.340060202) / PROTEINS-STRUCTURE FUNCTION AND GENETICS (1989)
  30. LAKOWICZ J.R. PRINCIPLES FLUORESCE (1983).
  31. 10.1126/science.3289117
  32. LEAR, J.D., SYNTHETIC AMPHIPHILIC PEPTIDE MODELS FOR PROTEIN ION CHANNELS, SCIENCE 240: 1177 (1988). (10.1126/science.2453923) / SCIENCE (1988)
  33. MARQUSEE, S, HELIX STABILIZATION BY GLU- ... LYS+ SALT BRIDGES IN SHORT PEPTIDES OF DENOVO DESIGN, PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA 84: 8898 (1987). / PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA (1987)
  34. Mathews, F. S., Cold Spring Harbor Symposia on Quantitative Biology 36: 387 (1972). (10.1101/SQB.1972.036.01.050) / Cold Spring Harbor Symposia on Quantitative Biology (1972)
  35. MATHEWS, F.S., STRUCTURE OF CYTOCHROME-B562 FROM ESCHERICHIA-COLI AT 2.5-A-RESOLUTION, JOURNAL OF BIOLOGICAL CHEMISTRY 254: 1699 (1979). (10.1016/S0021-9258(17)37829-8) / JOURNAL OF BIOLOGICAL CHEMISTRY (1979)
  36. MORIZE, I, REFINEMENT OF THE C2221 CRYSTAL FORM OF OXIDIZED UTEROGLOBIN AT 1.34-A RESOLUTION, JOURNAL OF MOLECULAR BIOLOGY 194: 725 (1987). (10.1016/0022-2836(87)90250-6) / JOURNAL OF MOLECULAR BIOLOGY (1987)
  37. MOSER, R, AN ARTIFICIAL CRYSTALLINE DDT-BINDING POLYPEPTIDE, FEBS LETTERS 157: 247 (1983). (10.1016/0014-5793(83)80555-9) / FEBS LETTERS (1983)
  38. MOSER, R, EXPRESSION OF THE SYNTHETIC GENE OF AN ARTIFICIAL DDT-BINDING POLYPEPTIDE IN ESCHERICHIA-COLI, PROTEIN ENGINEERING 1: 339 (1987). (10.1093/protein/1.4.339) / PROTEIN ENGINEERING (1987)
  39. MUTTER, M, THE CONSTRUCTION OF NEW PROTEINS .5. A TEMPLATE-ASSEMBLED SYNTHETIC PROTEIN (TASP) CONTAINING BOTH A 4-HELIX BUNDLE AND BETA-BARREL-LIKE STRUCTURE, PROTEINS-STRUCTURE FUNCTION AND GENETICS 5: 13 (1989). (10.1002/prot.340050104) / PROTEINS-STRUCTURE FUNCTION AND GENETICS (1989)
  40. MUTTER, M, NATURES RULES AND CHEMISTS TOOLS - A WAY FOR CREATING NOVEL PROTEINS, TRENDS IN BIOCHEMICAL SCIENCES 13: 260 (1988). (10.1016/0968-0004(88)90159-4) / TRENDS IN BIOCHEMICAL SCIENCES (1988)
  41. NAMBA, K, VISUALIZATION OF PROTEIN-NUCLEIC ACID INTERACTIONS IN A VIRUS - REFINED STRUCTURE OF INTACT TOBACCO MOSAIC-VIRUS AT 2.9-A RESOLUTION BY X-RAY FIBER DIFFRACTION, JOURNAL OF MOLECULAR BIOLOGY 208: 307 (1989). (10.1016/0022-2836(89)90391-4) / JOURNAL OF MOLECULAR BIOLOGY (1989)
  42. OHGUSHI, M, MOLTEN-GLOBULE STATE - A COMPACT FORM OF GLOBULAR-PROTEINS WITH MOBILE SIDE-CHAINS, FEBS LETTERS 164: 21 (1983). (10.1016/0014-5793(83)80010-6) / FEBS LETTERS (1983)
  43. 10.1126/science.2911757
  44. PACE, C.N., CRITICAL REV BIOCH 1: 1 (1975). / CRITICAL REV BIOCH (1975)
  45. PACE, C.N., METHODS IN ENZYMOLOGY 131: 266 (1986). (10.1016/0076-6879(86)31045-0) / METHODS IN ENZYMOLOGY (1986)
  46. PONDER, J.W., TERTIARY TEMPLATES FOR PROTEINS - USE OF PACKING CRITERIA IN THE ENUMERATION OF ALLOWED SEQUENCES FOR DIFFERENT STRUCTURAL CLASSES, JOURNAL OF MOLECULAR BIOLOGY 193: 775 (1987). (10.1016/0022-2836(87)90358-5) / JOURNAL OF MOLECULAR BIOLOGY (1987)
  47. 10.1126/science.2837824
  48. PTITSYN, O.B., THEORY OF PROTEIN SECONDARY STRUCTURE AND ALGORITHM OF ITS PREDICTION, BIOPOLYMERS 22: 15 (1983). (10.1002/bip.360220105) / BIOPOLYMERS (1983)
  49. PTITSYN, O.B., PROTEIN FOLDING - HYPOTHESES AND EXPERIMENTS, JOURNAL OF PROTEIN CHEMISTRY 6: 273 (1987). (10.1007/BF00248050) / JOURNAL OF PROTEIN CHEMISTRY (1987)
  50. 10.1126/science.3043666
  51. RICE, D.W., THE SPATIAL STRUCTURE OF HORSE SPLEEN APOFERRITIN, ADVANCES IN INORGANIC BIOCHEMISTRY 5: 39 (1983). / ADVANCES IN INORGANIC BIOCHEMISTRY (1983)
  52. RICHARDSON, J.S., ADVANCES IN PROTEIN CHEMISTRY 34: 167 (1981). (10.1016/S0065-3233(08)60520-3) / ADVANCES IN PROTEIN CHEMISTRY (1981)
  53. RICHARDSON, J.S., DENOVO DESIGN AND SYNTHESIS OF A PROTEIN, BIOPHYSICAL JOURNAL 45: A25 (1984). (10.1016/S0006-3495(84)84175-2) / BIOPHYSICAL JOURNAL (1984)
  54. RICHARDSON, J.S., EMBL BIOCOMPUTING TE 1: 71 (1986). / EMBL BIOCOMPUTING TE (1986)
  55. Richardson, J. S., Prediction of Protein Structure and Protein Conformation: 1 (1989). / Prediction of Protein Structure and Protein Conformation (1989)
  56. Richardson, J. S., Protein Engineering: 149 (1987). / Protein Engineering (1987)
  57. 10.1126/science.3381086
  58. RICHARDSON, J.S., THE DENOVO DESIGN OF PROTEIN STRUCTURES, TRENDS IN BIOCHEMICAL SCIENCES 14: 304 (1989). (10.1016/0968-0004(89)90070-4) / TRENDS IN BIOCHEMICAL SCIENCES (1989)
  59. ROSENBERG, A.H., VECTORS FOR SELECTIVE EXPRESSION OF CLONED DNAS BY T7 RNA-POLYMERASE, GENE 56: 125 (1987). (10.1016/0378-1119(87)90165-X) / GENE (1987)
  60. SAUER, R.T., AN ENGINEERED INTERSUBUNIT DISULFIDE ENHANCES THE STABILITY AND DNA-BINDING OF THE N-TERMINAL DOMAIN OF LAMBDA-REPRESSOR, BIOCHEMISTRY 25: 5992 (1986). (10.1021/bi00368a024) / BIOCHEMISTRY (1986)
  61. SHOEMAKER, K.R., TESTS OF THE HELIX DIPOLE MODEL FOR STABILIZATION OF ALPHA-HELICES, NATURE 326: 563 (1987). (10.1038/326563a0) / NATURE (1987)
  62. STENKAMP, R.E., STRUCTURE OF METHEMERYTHRIN AT 2.8-A RESOLUTION - COMPUTER GRAPHICS FIT OF AN AVERAGED ELECTRON-DENSITY MAP, BIOCHEMISTRY 17: 2499 (1978). (10.1021/bi00606a007) / BIOCHEMISTRY (1978)
  63. STUDIER, F.W., USE OF BACTERIOPHAGE-T7 RNA-POLYMERASE TO DIRECT SELECTIVE HIGH-LEVEL EXPRESSION OF CLONED GENES, JOURNAL OF MOLECULAR BIOLOGY 189: 113 (1986). (10.1016/0022-2836(86)90385-2) / JOURNAL OF MOLECULAR BIOLOGY (1986)
  64. UNSON, C.G., FEDERATION PROCEEDINGS 43: 1837 (1984). / FEDERATION PROCEEDINGS (1984)
  65. WEBER, P.C., STRUCTURE OF CYTOCHROME-C' - A DIMERIC, HIGH-SPIN HEME PROTEIN, NATURE 286: 302 (1980). (10.1038/286302a0) / NATURE (1980)
Dates
Type When
Created 18 years, 10 months ago (Oct. 5, 2006, 6:10 p.m.)
Deposited 1 year, 7 months ago (Jan. 12, 2024, 11:19 a.m.)
Indexed 2 months ago (June 26, 2025, 9:10 a.m.)
Issued 35 years ago (Aug. 24, 1990)
Published 35 years ago (Aug. 24, 1990)
Published Print 35 years ago (Aug. 24, 1990)
Funders 0

None

@article{Hecht_1990, title={De Novo Design, Expression, and Characterization of Felix: a Four-Helix Bundle Protein of Native-Like Sequence}, volume={249}, ISSN={1095-9203}, url={http://dx.doi.org/10.1126/science.2392678}, DOI={10.1126/science.2392678}, number={4971}, journal={Science}, publisher={American Association for the Advancement of Science (AAAS)}, author={Hecht, Michael H. and Richardson, Jane S. and Richardson, David C. and Ogden, Richard C.}, year={1990}, month=aug, pages={884–891} }