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3D Structures of Biological Macromolecules Part 1: Introduction - Facts, Tools and Fascination Jürgen Sühnel [email protected] Leibniz Institute for Age Research, Fritz Lipmann Institute, Jena Centre for Bioinformatics Jena / Germany Supplementary Material: www.fli-leibniz.de/www_bioc/3D/ Fritz Lipmann The Journal of Biological Chemistry 186, 235, 1950 Coenzyme A Computational Structural Biology Biocomputing Group: Research and Service Identification and Analysis of Unusual Structural Motifs in Proteins and Nucleic Acids • Water-Mediated Base Pairs • Base Polyad Motifs in DNA Mutiplex Structures • C-H...O/N Interactions in RNA Structures • C-H... Interactions in Proteins (IMB, X-Ray Crystallography: Weiss) • Nest Motifs and Non-repetitive Dipeptide Patterns in Proteins Other Computational Genomics (UCLA: Pal; EMBL Hamburg Outstation: Weiss) Database ‚Jena Library of Biological Macromolecules (JenaLib)‘ Computational Genomics of Prokaryotes (IMB/FLI Genome Analysis Group: Platzer) Short-term Collaborative Projects • Structure and Conformational Analysis of Ampullosporin A (HKI: Gräfe; Friedrich Schiller University: Görls, Reissmann) • Computer Selection of DNA Oligonucleotide Probe Combinations for COMBO-FISH • Chromatin Structure (Heidelberg/Freiburg University: Hausmann) (FLI, Molecular Biology Group: Diekmann) Websites for Molecular Biology Service • The RNA World Website • The JCB Protein-Protein Interaction Website Maintenance of the Institutional Computer Network www.fli-leibniz.de/www_bioc/ Biocomputing Group: Databases, Websites/Webtools The Jena Library of Biological Macromolecules (JenaLib) www.fli-leibniz.de/IMAGE.html The RNA World Website www.fli-leibniz.de/RNA.html The JCB Protein-Protein Interaction Website www.fli-leibniz.de/jcb/ The Jena Prokaryotic Genome Viewer jpgv.fli-leibniz.de The Spirochetes Genome Browser sgb.fli-leibniz.de Biocomputing Group: Methods Quantum Chemistry Molecular Dynamics (Structural) Bioinformatics Computational Genomics Database / Website Development Different Protein Views Structural Biology Genomics Porin (1a0s) Cell Biology Systems Biology SIR2p – NAD+-dependent histone deacetylase PNC1 – pyrazinamidase/nicotinamidase 1) Internal Co-ordinates (Bond) Distance (Bond) Angle Torsion angle [2 atoms] [3 atoms] [4 atoms] C D B A Protein Structure: Amino Acids Protein Structure: Amino Acids www.fli-leibniz.de/IMAGE_AA.html Protein Structure Sequence Protein-protein or protein-nucleic acid complexes Secondary structure elements Domains/Folds/Chains Protein Structure: Secondary Structure Elements Protein Structure: The Protein -Helix The first and still one of the greatest triumphs of speculative model building in structural chemistry/biology Forerunner of computer-assisted model building In the PNAS paper the helix is drawn left-handed !!!!! Linus Pauling and Robert Corey with a wooden model of a protein -helix (1 inch per Å) Pauling, Corey, Branson. The structure of proteins: Two hydrogen-bonded helical conformations of the polypeptide chain. Proc. Natl. Acad. USA 1951, 37, 205-211. Protein Structure: Domains Structure-based definition Dali Fold Classification Sequence-based definition Protein Domains: Jenalib Jmol Viewer calcium-free human calpain Calpains (calcium-dependent cytoplasmic cysteine proteinases) are implicated in processes such as cytoskeleton remodeling and signal transduction. Protein Domains: JenaLib Jmol Viewer Protein Structure: Backbone Torsion Angles D. W. Mount: Bioinformatics, Cold Spring Harbor Laboratory Press, 2001. Protein Structure: The Ramachandran Map Theoretical Beta-sheet Left-handed alpha-helix Right-handed alpha-helix Ramachandran, Ramakrishnan, Sasisekharan. Stereochemistry and polypeptide chain configurations. J. Mol. Biol. 1963, 7, 95-99. Ramachandran, Sasisekheran. Conformation of polypeptides and proteins. Adv. Protein. Chem. 1968, 23, 283-438. Experimental 237 384 amino acids in 1041 protein chains Hovmöller et al., Acta Cryst. D 2002, 58, 768-776 non-glycines glycines Protein Structure: History Myoglobin Model Built by A. A. Barker, Model Maker in Cambridge (UK) www.umass.edu/molvis/francoeur/barker/barker.html Protein Structure: Hemoglobin Model www2.mrc-lmb.cam.ac.uk/archive/Perutz62.html The Oldest Myoglobin and Hemoglobin Structures in the PDB Myoglobin (1mbn) Hemoglobin (2dhb) 2 alpha chains: 146 aa (green, cyan) 2 beta chains: 141 aa (red, brown) (Watson, Kendrew: 1973) (Perutz et al.: 1973) Protein Structure: Thermodynamic Hypothesis Christian B. Anfinsen The native structure of a protein corresponds to the minimum of free energy. Protein Structure: Free energy The protein in solution is viewed as a statistical ensemble. G = H - T S G - Free energy H - Enthalpy S - Entropy The enthalpy is governed by all interactions in a molecular system. The entropy is governed by the degrees of freedom. H can, in principle, be determined from quantum-mechanical/chemical calculations. S can be determined by statistical-mechanical methods. Often a simplified description of the forces/potentials/interaction energies that govern protein structure is used. Protein Structure Simplified description of interactions (forces) governing protein structure Covalent interactions Covalent bonds including SS-interactions Non-covalent interactions Hydrogen bonds Electrostatic interactions Salt bridges Van-der-Waals interactions Cation--interactions - interactions (stacking) C-H…O/N and C-H… interactions N/O-H… interactions The hydrophobic effect More Hydrogen Bonds for the (Structural) Biologist Nucleic Acid Structure Nucleic Acid Structure: Nucleotide Nucleic acids are polymers made up of repeated units, nucleotides, comprising three components: - phosphate, - a sugar (2'-deoxyribose in DNA, ribose in RNA), - and one of four heretocyclic bases. In a formal sense a nucleic acid strand is generated by forming C3'-O3' bonds between different nucleotides. This is, however, only a formal structural description. The chemical reaction is more complicated. The well-known double helix is obtained by connecting the two strands via hydrogen bonding between bases. www.fli-leibniz.de/IMAGE_BPDIR.html Nucleic Acid Structure: Bases www.fli-leibniz.de/IMAGE_BPDIR.html DNA Structure: History DNA Structure: History DNA Structure: Ideal B-DNA Conformation First Single-Crystal DNA Structure (B-DNA) Drew-Dickerson structure H. R. Drew, R. M. Wing, T. Takano, C. Broka, S. Tanaka, K. Itakura, R. E. Dickerson Structure Of A B-/DNA Dodecamer. Conformation And Dynamics Proc. Nat. Acad. Sci. Usa V. 78 2179 1981 The Very First 3D Biopolymer Structures The first three-dimensional structure of a biopolymer was the DNA model built by J. D. Watson and F. H. C. Crick in 1953 taking into account fiber diffraction data provided by M. H. F. Wilkins and others (Nobel Prize in Physiology or Medicine, 1962). The very first single-crystal DNA structure was resolved by Dickerson and co-workers (PDB code: 1bna). The first three-dimensional protein structures (myoglobin and hemoglobin) were determined by M. F. Perutz and J. C. Kendrew (Nobel Prize in Chemistry, 1962). The entries included in the PDB (PDB codes: 1mbn and 2dhb) represent refined structures. J. C. Kendrew had obtained a myoglobin structure at a resolution of 6 Å already in 1957. A. Klug has contributed substantially to the development of electron microscopy. This method is suitable for systems that cannot be crystallized (Nobel Prize in Chemistry, 1982). A. Klug has applied this method primarily to virus structures (see for example, Finch, Klug, J. Mol. Biol. 1965, 13, 1-12). The structure of the first membrane-bound protein (a photosynthetic reaction centre; PDB code: 1prc) was resolved by J. Deisenhofer, R. Huber and H. Michel (Nobel Prize in Chemistry, 1988). The first three-dimensional protein structure determined by NMR spectroscopy was proteinase IIa inhibitor from bull seminal plasma (PDB codes: 1bus, 2bus). It was reported by K. Wüthrich and co-workers in 1985 (Nobel Prize in Chemistry, 2002). Note, that in the early days of structural biology the PDB did not exist. It was created in 1970-1971. Databases with 3D Structural Information Comprehensive resources of biological macromolecules Protein Data Bank (PDB) PDBsum Macromolecular Structure Data Base (MSD) OCA Jena Library of Biological Macromolecules (JenaLib) Nucleic-acid containing structures only Nucleic Acid Database (NDB) RNA Structure Database (RNAbase) Protein structures only Database of Comparative Protein Structure Models - ModBase Structure-based classification of proteins CATH SCOP Small molecules National Cancer Institute 3D structure database (collection of 3D structures for over 400,000 drugs) Cambridge Structural Database (CSD) (organic and metal-organic compounds including polypeptides and polysaccharides up to 24 units) For weblinks see: www.fli-leibniz.de/www_bioc/3D/ PDB File HEADER HYDROLASE (ENDORIBONUCLEASE) 12-MAY-95 1RTU TITLE USTILAGO SPHAEROGENA RIBONUCLEASE U2 COMPND MOL_ID: 1; COMPND 2 MOLECULE: RIBONUCLEASE U2; COMPND 3 CHAIN: NULL SOURCE MOL_ID: 1; SOURCE 2 ORGANISM_SCIENTIFIC: USTILAGO SPHAEROGENA; SOURCE 3 ORGANISM_COMMON: SMUT FUNGUS; SOURCE 4 ATCC: 12421 KEYWDS HYDROLASE, ENDORIBONUCLEASE, BETA-ISOMERIZED ASPARTATE EXPDTA X-RAY DIFFRACTION AUTHOR S.NOGUCHI,Y.SATOW,T.UCHIDA,C.SASAKI,T.MATSUZAKI REVDAT 1 08-NOV-96 1RTU 0 JRNL AUTH S.NOGUCHI,Y.SATOW,T.UCHIDA,C.SASAKI,T.MATSUZAKI JRNL TITL CRYSTAL STRUCTURE OF USTILAGO SPHAEROGENA JRNL TITL 2 RIBONUCLEASE U2 AT 1.8 A RESOLUTION JRNL REF BIOCHEMISTRY V. 34 15583 1995 JRNL REFN ASTM BICHAW US ISSN 0006-2960 0033 REMARK 1 REMARK 2 REMARK 2 RESOLUTION. 1.80 ANGSTROMS. REMARK 3 REMARK 3 REFINEMENT. REMARK 3 PROGRAM : PROLSQ REMARK 3 AUTHORS : KONNERT,HENDRICKSON . . . SEQRES 1 114 CYS ASP ILE PRO GLN SER THR ASN CYS GLY GLY ASN VAL SEQRES 2 114 TYR SER ASN ASP ASP ILE ASN THR ALA ILE GLN GLY ALA SEQRES 3 114 LEU ASP ASP VAL ALA ASN GLY ASP ARG PRO ASP ASN TYR SEQRES 4 114 PRO HIS GLN TYR TYR IAS GLU ALA SER GLU ASP ILE THR SEQRES 5 114 LEU CYS CYS GLY SER GLY PRO TRP SER GLU PHE PRO LEU SEQRES 6 114 VAL TYR ASN GLY PRO TYR TYR SER SER ARG ASP ASN TYR SEQRES 7 114 VAL SER PRO GLY PRO ASP ARG VAL ILE TYR GLN THR ASN SEQRES 8 114 THR GLY GLU PHE CYS ALA THR VAL THR HIS THR GLY ALA SEQRES 9 114 ALA SER TYR ASP GLY PHE THR GLN CYS SER MODRES 1RTU IAS 45 ASP BETA CARBOXY LINKED TO NEXT RESIDUE . ATOM 1 N CYS 1 -4.933 19.344 18.255 1.00 5.83 ATOM 2 CA CYS 1 -6.309 19.098 17.823 1.00 7.72 ATOM 3 C CYS 1 -6.270 18.283 16.532 1.00 8.17 ATOM 4 O CYS 1 -5.179 17.995 16.022 1.00 8.08 ATOM 5 CB CYS 1 -7.128 20.382 17.691 1.00 6.94 ATOM 6 SG CYS 1 -6.599 21.528 16.386 1.00 8.61 ATOM 7 N ASP 2 -7.451 17.930 16.012 1.00 9.26 . N C C O C S N Analysis and Modeling Tools • Web-based tools • Standalone programs See compilation at www.fli-leibniz.de/www_bioc/3D/ Cross references from other databases Databases RNABase Systers ~ 1 information request per minute Reichert, Sühnel. The IMB Jena Image Library of Biological Macromolecules - 2002 update. Nucleic Acids Res. 2002, 30, 253-254. Bioinformatics Group (K. Schmid) PDB Content Growth 1727 structures (~ 2 structures per day) = 24516 structures (~13 structures per day) = 29841 structures (~15 structures per day) Start 1993: 2003: 23792 + 724 2004: 28992 + 849 PDB Content Statistics Total number of entries: 36344 (1063) Entries listed by molecule type: Proteins Protein-nucleic acid complexes Nucleic acids Other (Carbohydrates, …) 33223 (945) 1492 (63) 1595 (55) 34 (0) Entries listed by method of structure determination: Diffraction NMR Electron Microscopy Other Modeling 30784 5361 123 76 1063 May 2, 2006 PDB Extremes Entry with longest chain: 1ofd: Glutamate synthase [2 chains with 1491/1507 amino acids each; each chain consists of 3 domains according to SCOP] Glutamate synthases (GltS) are crucial enzymes in ammonia assimilation in plants and bacteria, where they catalyze the formation of two molecules of L-glutamate from L-glutamine and 2-oxoglutarate. The plant-type ferredoxin-dependent GltS and the functionally homologous alpha subunit of the bacterial NADPH-dependent GltS are complex four-domain monomeric enzymes of 140-165 kDa belonging to the NH(2)-terminal nucleophile family of amidotransferases. The enzymes function through the channeling of ammonia from the N-terminal amidotransferase domain to the FMN-binding domain. Entries with largest number of chains: 1otz/1p0t Baff-baff-R complex [60 chains, two PDB files required] B-cell activating factor (BAFF) is a key regulator of B-lymphocyte development. Its biological role is mediated by the specific receptors BCMA, TACI and BAFF-R. The cysteine-rich domain (CRD) of the BAFF-R extracellular domain adopts a beta-hairpin structure and binds to the virus-like BAFF cage in a 1:1 molar ratio. Entry with largest number of models: 1e9t: Human intestinal trefoil factor [NMR structure with 85 models] Human intestinal trefoil factor, hITF, a secretory polypeptide found mainly in the human gastrointestinal tract, is a member of the newly characterized trefoil factor or P-domain peptide family representing putative growth factors. PDB Extremes: Small Peptide (Peptaibol Alamethicin, chain A, 1amt) Three structural characteristics define the group of polypeptides known as peptaibols: A short chain length, typically between 15 and 20 residues, although shorter ones are known. A high proportion of the amino acid residues are non-standard. There is a particular propensity for Aminoisobutyric acid (Aib). The chain has an alkyl N terminus (usually acetyl) and a hydroxy-amino acid at the C terminus. Peptaibols generally exhibit antimicrobial activity and are referred to as antibiotic peptides. The main sources of the peptaibols known to date are fungii of the genre Trichoderma and Emericellopsis The antomicrobial activity is thought to arise from the peptaibols' membrane activity and their ability to form pores in lipid membranes. The pores so formed are able to conduct ionic species; this conductance leads to the loss of osmotic balance and cell death. Along with Aib, non standard residues which have been observed include Isovaline (Iva), Hydroxyproline (Hyp) and Ethylnorvaline (Etnor). Aminoisobutyric acid has a high tendency to form helices and this is borne out by the helical structures of the peptaibols. www.cryst.bbk.ac.uk/peptaibol/introduction.htm Quantitative helix bending analysis described in: Kronen et al., J. Peptide Science 2003, 9, 729-724 Crystal structure and conformational analysis of ampullosporin A BiologicalUnit Glutamate Synthase (1ofd) – JenaLib Atlas Page: Astex Viewer Glutamate Synthase (1ofd) – JenaLib Atlas Page: WebMol Viewer Glutamate Synthase (1ofd) – JenaLib Atlas Page: Fe3S4 Environment Baff-Baff-R complex (1otz/1p0t) This complex has 60 chains and therefore two PDB files are required for the complete structure, Vision www2.mrc-lmb.cam.ac.uk/groups/GS/eye.html Rhodopsin (1f88) - 2000 Rhodopsin (1f88) Microtubules Cytoskeleton of a cultured epithelial cell. Mircotubules are shown in green, actin is shown in red and DNA is blue. Image by Steve Rodgers. 3D Microtubule Image Reconstruction Microtubule Model sun.menloschool.org/~birchler/cells/animals/cytoskeleton/ www-vis.lbl.gov/Vignettes/KDowning-Microtubules/ users.rcn.com/jkimball.ma.ultranet/BiologyPages/C/Cytoskeleton.html b-Tubulin Dimer Complexed with a Taxol Derivative (1tub) - 1997 The structure was solved by electron microscopy. Ramachandran Maps of the Tubulin/Taxol Complex Original structure at a resolution of 3.9 Å. Refined structure at a resolution of 3.5 Å. E. Nogales, S. G. Wolf, K. H. Downing Structure Of The Alpha Beta Tubulin Dimer By Electron Crystallography Nature V. 391 199 1998 J. Lowe, H. Li, K. H. Downing, E. Nogales Refined Structure Of Alpha-Beta Tubulin At 3. 5 A Resolution J. Mol. Biol. V. 313 1045 2001 Taxol (Paclitaxel) Pacific yew tree Taxus brevifolia A happy team of FSU Taxol researchers led by Bob Holton (bottom row, right) claimed victory in totally synthesizing the drug on Dec. 9, 1993. Hope in a Bottle: Bristol-Myers Squibb introduced Taxol to the marketplace in January 1993. From bark to business, the process took 31 years. www.research.fsu.edu/researchr/fall2002/taxol.html Cover Images for all 2003/2004/2005/2006 Issues of the Journal RNA t-RNA (4tna) - 1978 Large Ribosomal Subunit (1jj2) - 2001 Nucleosome Core Particle (1aoi) - 1997 DNA in chromatin is organized in arrays of nucleosomes.Two copies of each histone protein, H2A, H2B, H3 and H4, are assembled into an octamer that has 145-147 base pairs of DNA wrapped around it to form a nucleosome core. Jena Structures: 5S rRNA E-loop/L25 Complex (1d6k) - 1999 IMB/FLI: Molecular Biophysics/NMR Spectroscopy (M. Görlach) Jena Structures: Macrophage Infectivity Potentiator Protein (1fd9) - 2001 The human pathogen Legionella pneumophila, the etiological agent of the severe and often fatal Legionnaires' disease, produces a major virulence factor, termed 'macrophage infectivity potentiator protein' (Mip), that is necessary for optimal multiplication of the bacteria within human alveolar macrophages. Mip exhibits a peptidyl prolyl cis-trans isomerase (PPIase) activity, which appears to be important for infection. IMB/FLI: Structural Biology / Crystallography (R. Hilgenfeld) Integration Host Factor (1ihf) - 1996 An architectural protein which assists formation of high order protein-DNA complexes such as those found in replication and long distance transcription regulation. Accelrys WebLab Viewer See www.imb-jena.de/www_bioc/3D/ for availability RasMol See www.imb-jena.de/www_bioc/3D/ for availability Program Output of the Program secstr (Part of PROMOTIF) PDB numbering Continous numbering phi psi secondary structure assignment omega Newspapers Gallery Gallery