Research Report 2000 - MDC

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Folding and Misfolding of Proteins Gregor Damaschun The creation of proteins in living cells consists of two main processes: biosynthesis of the polypeptide chain and its folding into the native, threedimensional structure with biological function. The first process has been thoroughly studied, while the second process is less well known. We have learnt in recent years that the proteinfolding process is not always flawless within the cell and this can have pathological consequences. Thus, a number of human diseases are related to the deposition of protein fibrils causing tissue damage and degeneration. Amyloid fibrils develop from abnormal, misfolded conformational states of different normally soluble proteins forming ordered aggregates. The reasons for misfolding are unknown. Therefore, there are no causal treatments for these diseases. The group “Physics of Biopolymers” is engaged in studies of the folding pathways of proteins to understand the causes of misfolding. The main experimental methods used in these studies include solution X-ray scattering (SOXS), dynamic light scattering (DLS) and optical spectroscopy, including kinetic techniques. Methods of statistical physics of chain molecules have been applied to modelling the experimental data. 46 Polymorphism of proteins Textbooks state that the structure of a protein is determined by its amino acid sequence. However, we have been able to show experimentally that this so-called second genetic code is not unambiguous. The threedimensional structure of a protein is determined not only by the amino acid sequence but also by the environment of the protein molecules and is influenced by interactions between structural intermediates on the folding pathway. Therefore, many proteins can adopt differently folded threedimensional structures and only one of these structures is functionally active. For yeast phosphoglycerate kinase (PGK), we observed in addition to the native structure two further, different conformations. The starting point for the formation of these misfolded conformations is the acid-unfolded state. At low pH values, PGK has the conformation of an expanded random walk. If the molecule is transferred to a hydrophobic environment with a low dielectric constant, the entire molecule forms α-helix. On the other hand, anion-induced partial refolding of the acid-unfolded state leads to the formation of amyloid-like fibrils. Half the amino acids have the conformation of a cross-β-helix which is typical of all amyloids. Folding pathways and kinetics The formation of amyloids starts from non-natively folded monomeric intermediates. The monomers aggregate forming successively dimers, tetramers and octamers. More and more cross-β-structure develops during this aggregation process. The kinetics of aggregation is strongly dependent on protein concentration. At room temperature, this process may take several hours. Subsequently, the octamers grow in one direction only and form fibrils. The growth of the fibrils, i.e. their time-dependent elongation, may take some months. Our results indicate that inhibitors of cross-β-structure formation can be effective only during the early phases of amyloidosis. The slow kinetics are typical of misfolding of proteins into amyloids. In vivo, the progression of these processes is in some cases even slower than in our in vitro experiments. By contrast, the folding of a protein into its native structure is a fast process. Typical times for folding vary from milliseconds to minutes. One central problem in protein folding is the question, whether chain segments with a periodic secondary structure develop in a first step, then form in a second step the compact globule through diffusion (framework model), or whether the chain initially collapses, driven by hydrophobic interactions, with concurrent or subsequent formation of segments with periodic secondary structure (hydrophobic collapse model). We have been able to show experimentally that both models are not general alternatives. There are proteins folding mainly according to the mechanism of the framework model (e.g., bovine RNase A) as well as folding according to the hydrophobic collapse model (e.g., bovine α-lactalbumin). Further studies are necessary to address the open question: which of these folding scenarios is more prone to the misfoldings that lead to amyloids? Up to now, a search for common properties of amyloid-forming proteins has been unsuccessful.
Selected Publications Damaschun, G., Damaschun, H., Gast, K., and Zirwer, D. (1998) Denatured states of yeast phosphoglycerate kinase. Biochemistry (Moscow) 63, 259-275. Gast, K., Zirwer, D., Müller-Frohne, M., and Damaschun, G. (1998) Compactness of the kinetic molten globule of bovine α-lactalbumin: A dynamic light scattering study. Protein Sci. 7, 2004-2011. Nöppert, A., Gast, K., Zirwer, D., and Damaschun, G. (1998) Initial hydrophobic collapse is not necessary for folding RNase A. Fold. Des. 3, 213-221. Gast, K., Zirwer, D., Müller-Frohne, M., and Damaschun, G. (1999) Triflouroethanol-induced conformational transition of proteins: insights gained from the differences between α-lactalbumin and ribonuclease A. Protein Sci. 8, 625-634. Damaschun, G., Damaschun, H., Gast, K., and Zirwer, D. (1999) Proteins can adopt totally different folded conformations. J. Mol. Biol. 291, 715- 725. Figure 22: Formation of amyloid fibrils by misfolding of proteins. The blue bars represent cross-β-structures of the polypeptide chain. Uunfolded state in an acidic environment, Nnative state, I-folding intermediate. U N I Structure of the Group Group leader Prof. Dr. Gregor Damaschun Scientists Hilde Damaschun Dr. Klaus Gast Dr. Dietrich Zirwer Graduate and undergraduate students Ansgar Siemer Technical assistant Reinhard Kröber Amyloid 47
Page 1 and 2: Research Report 2000 Covers the Per
Page 3 and 4: Molecular Therapy Molecular and Dev
Page 5 and 6: The quantum theory, that provided a
Page 7 and 8: Introduction Clinical Research The
Page 9 and 10: Genome Research in Berlin- Brandenb
Page 11 and 12: External Evaluation Over the period
Page 13 and 14: Congresses In the years reported, t
Page 15 and 16: Awards A number of prestigious priz
Page 17 and 18: Genetics, Bioinformatics and Struct
Page 19 and 20: disorders. Various MDC groups have
Page 21 and 22: Steen R.G., Kwitek-Black A.E., Glen
Page 23 and 24: Selected Publications Binas, B., Da
Page 25 and 26: Technology transfer Based on the fu
Page 27 and 28: Selected Publications Müller, D.N.
Page 29 and 30: Selected Publications Hennies, H.C.
Page 31 and 32: Somatic genetic alterations in brea
Page 33 and 34: Clinical and Molecular Genetics of
Page 35 and 36: Selected Publications Toka, H.R., B
Page 37 and 38: Lbx1, c-met and the control of cell
Page 39 and 40: Selected Publications Moore, A., Mc
Page 41 and 42: Selected Publications Herz, J., Wil
Page 43 and 44: We aim to study the genetic epidemi
Page 45: Nucleation Nucleation as a pre-requ
Page 49 and 50: Selected Publications Yuan, T., Wal
Page 51 and 52: stability. By determining the struc
Page 53 and 54: Role of Protein Dynamics in Enzyme
Page 55 and 56: Modeling Nucleic Acid Structure and
Page 57 and 58: Conformation, Stability and Interac
Page 59 and 60: Protein Structure Analysis and Prot
Page 61 and 62: Cell Growth and Differentiation 61
Page 63 and 64: dendritic cells (DC). Adoptive tran
Page 65 and 66: developmental pathway that may invo
Page 67 and 68: Regulation of Transcription in Mamm
Page 69 and 70: Differentiation and Growth Control
Page 71 and 72: Cell cycle-dependent transcriptiona
Page 73 and 74: Cell Cycle Regulation Hans-Dieter R
Page 75 and 76: Epithelial Differentiation, Invasio
Page 77 and 78: Selected Publications Hartmann, G.,
Page 79 and 80: Selected Publications Behrens, J.,
Page 81 and 82: Selected Publications Karsten, U.,
Page 83 and 84: Depressed contractility in human he
Page 85 and 86: Understanding calcium handling prot
Page 87 and 88: VSMCs. We have cloned and character
Page 89 and 90: Surgical Oncology Peter M. Schlag T
Page 91 and 92: Ubiquitin System and Endoplasmic Re
Page 93 and 94: P450 Cytochromes and the Endoplasmi
Page 95 and 96: Vascular Biology Hermann Haller Thi
Page 97 and 98:
Selected Publications Gollasch, M.,
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Electron Microscopy Members of the
Page 101 and 102:
Molecular Therapy 101
Page 103 and 104:
Hematology, Oncology and Tumor Immu
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Selected Publications Sturm, I., K
Page 107 and 108:
Selected Publications Westermann, J
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fractions. The hematopoietic potent
Page 111 and 112:
Interaction of pharmacologically ac
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Molecular Basis of Congestive Heart
Page 115 and 116:
Selected Publications Schneider, G.
Page 117 and 118:
different murine and human tumor ce
Page 119 and 120:
Molecular and Cell Biology of Hemat
Page 121 and 122:
Phospholipids Dietrich Arndt Cytoto
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Regulation and Deregulation of Cell
Page 125 and 126:
Evolution, Regulation and Genetic A
Page 127 and 128:
Molecular and Developmental Neurosc
Page 129 and 130:
Cellular Neurosciences Helmut Kette
Page 131 and 132:
Growth Factor and Regeneration Gary
Page 133 and 134:
Synapse Formation and Function Fran
Page 135 and 136:
Developmental Neurobiology Fritz G.
Page 137 and 138:
Selected Publications Volkmer, H.,
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Structure and Organization 139
Page 141 and 142:
Prof. Dr. Hans Meyer President of t
Page 143 and 144:
Supporting Divisions Safety The div
Page 145 and 146:
Press and Public Relations Research
Page 147 and 148:
Purchasing and Materials Management
Page 149 and 150:
Computing The computing group of th
Page 151 and 152:
Awards Thomas Biederer Boehringer-M
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Index A Abdul, Yetunde 90 Aguirre-A
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H Haase, Hannelore 85, 97, 100, 142
Page 157 and 158:
Q Qin, Zhihai 107, 117 Quass, Petra
Page 159:
Board of Trustees Chair Wolf-Michae
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Research Report 2000 - MDC

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