Protein Classification and Structure Prediction Amino acid ...

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Levels of Protein Structure • Secondary structure elements combine to form tertiary structure • Quaternary structure occurs in multi-enzyme complexes • Many proteins are active only as homodimers, homotetramers, , etc. Protein Structure Examples Krane & Raymer • Two cysteines in close proximity will form a covalent bond • Disulfide bond, disulfide bridge, or dicysteine bond. • Significantly stabilizes tertiary structure. Disulfide Bonds Determining Protein Structure • There are O(100,000) distinct proteins in human proteome. • Two methods for revealing positions of atoms in 3-D: 3 • X-Ray Crystallography • X-ray diffraction pattern + mathematical construction • Good protein crystal needed, good resolution of diffraction needed ed • Nuclear Magnetic Resonance • Small proteins only (< 250 residues) • Inter-proton distances + geometric constraints • 24,000+ 3D structures have been determined so far (including duplicates with different ligand bound, etc.) Krane & Raymer PDB Holdings List: 23-Mar Mar-2004 PDB: Growth (Mar 2004) X-ray Diffraction and other NMR Proteins, Peptides, & Viruses 19427 2998 Protein/Nucleic Acid Complexes 944 97 Nucleic Acids 726 575 Carbohydrates 14 4 total 21111 3674 Total 22425 1041 1301 18 24785 Please note that theoretical models have been removed. 4
Experimentation to grow protein crystals X-Ray Crystallography Trial-and-error Experimentation Observables Partial Success ~0.5mm Trial 1 Control Parameters Failure Success Trial 3 Trial 2 • The crystal is a mosaic of millions of copies of the protein. • As much as 70% is solvent (water)! • May take months (and a “green” thumb) to grow. Krane & Raymer Krane & Raymer X-Ray diffraction • Image is averaged over: • Space (many copies) • Time (of the diffraction experiment) Protein Crystal Growth in Space • Protein crystal growth experiments on over 20 shuttle missions since 1984 • Larger Crystals in 45.4% of the cases • New Crystal Structures in 18% of the cases • ≥ 10% increase in X-Ray X Crystallography Brightness in 58% of the cases • Less thermal motion in 27.2% of the cases • An X-Ray X Crystallography resolution improvement of ~0.3 Å in 42.4% of the cases • An X-Ray X Crystallography resolution improvement of 0.3 to 0.5 Å in 9.9% of the cases • An X-Ray X Crystallography resolution improvement of 0.5 to 1.0 Å in 9.9% of the cases Krane & Raymer The Protein Folding Problem • Proteins self-assemble in solution. Almost all of the information necessary to determine the complex 3-D 3 structure is in the amino acid sequences • Central dogma: Sequence specifies structure • Central question: “Given a particular sequence of amino acid residues (primary structure), what will the tertiary/quaternary structure of the resulting protein be?” Levinthal’s paradox • Consider a 100 residue protein. If each residue can take only 3 positions, there are 3 100 = 5 × 10 47 possible conformations. • If it takes 10 -13 s to convert from 1 structure to another, exhaustive search would take 1.6 × 10 27 years! 5
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