臺灣博碩士論文加值系統

離子作用力對於穩定蛋白質結構及核糖核酸辨識扮演相當重要的角色，主要由帶相反電荷的胺基酸所形成。自然界中帶電荷胺基酸具有不同的側鏈長度，例如負電荷胺基酸中，Asp及Glu側鏈分別具有一個與兩個疏水性亞甲基團，非自然界負電荷胺基酸Aad則具有三個亞甲基團。為研究負電荷胺基酸側鏈長度對於β-hairpin結構穩定中之離子作用力的影響，遂設計合成出一系列帶有不同負電荷胺基酸Zbb (Asp, Glu及Aad)之β-hairpin胜肽 (HPTZbbArg)。利用二維核磁共振光譜可確認胜肽HPTAspArg、HPTGluArg及HPTAadArg所形成之β-hairpin結構。比較所合成之胜肽與控制組胜肽中各殘基α氫原子化學位移差異可計算出胜肽分子之摺疊比例與自由能。β-Hairpin跨股側向離子作用力之能量大小以雙突變循環分析，結果顯示：Aad-Arg > Glu-Arg ≈ Asp-Arg，可知側鏈愈長的負電荷胺基酸，其跨股側向離子作用力之能量愈大。
人類免疫缺陷病毒Tat蛋白中47-57區域對於專一性辨識TAR RNA及穿透細胞非常重要。Tat蛋白與TAR RNA之結合對於人類免疫缺陷病毒基因組複製是不可或缺的。為研究Arg雙甲基化對於TAR RNA辨識及穿透細胞之影響，將Tat蛋白中47-57區域之六個Arg分別取代成ADMA(不對稱雙甲基化)及SDMA(對稱雙甲基化)，利用膠體電泳位移分析Tat衍生物對TAR RNA之結合能力與專一性，並利用流式細胞儀評估Tat衍生物穿透Jurkat細胞膜且進入細胞內部的效率。

Ion pairing interactions play important roles in protein stability and RNA recognition. Ion pairs are formed between a pair of oppositely charged amino acids. Interestingly, natural charged amino acids have different number of hydrophobic methylenes on their side chains. For negatively charged residues, Asp has one methylene and Glu has two methylenes. The analogous non-encoded negatively charged amino acid, Aad, contains three methylenes. To study the effect of negatively charged amino acid side chain length on cross strand ion pairs in β-sheets stability, a basic β-hairpin model HPTZbbArg was designed. Zbb denotes the negatively charged residues. The hairpin structure for peptides HPTAspArg, HPTGluArg, and HPTAadArg were confirmed by NMR methods. The fraction folded of the peptides was determined using chemical shift data involving the fully unfolded and the fully folded reference peptides. The interaction free energy followed the trend: Aad-Arg > Glu-Arg ≈ AspArg. Apparently, the longer the negatively charged residue side chain length, the stronger the ion pairing interaction.
HIV-1 Tat protein contains an arginine-rich sequence (Tat49-57), which binds specifically to the trans-activating responsive (TAR) element and plays an important role in nuclear localization. The binding of HIV-1 Tat protein and TAR RNA is essential for HIV-1 virus genome replication. To study the effect of arginine dimethylation on RNA recognition and cellular uptake, each arginine residue in Tat49-57 was replaced with a dimethylated arginine including ADMA and SDMA, the asymmetric and symmetric dimethylated forms, respectively. The dissociation constant for the Tat derived peptide-TAR RNA complexes was determined by gel shift assay. The cellular uptake efficiency of these Tat derived peptides into Jurkat cells was assessed by flow cytometry.

誌謝---------------------------------------------------- i
中文摘要-------------------------------------------------ii
Abstract-----------------------------------------------iii
Table of Contents----------------------------------------v
List of Figures---------------------------------------viii
List of Tables------------------------------------------xi
Abbreviation------------------------------------------xiii
Chapter 1. Introduction----------------------------------1
1-1 The three Macromolecules: DNA, RNA, and Proteins-----1
1-2 The Roles of Protein Molecules-----------------------2
1-3 Protein Folding and Function-------------------------3
1-4 Hierarchy of Protein Structure-----------------------4
1-5 Dominant Forces for Protein Folding-----------------10
1-6 Salt-bridges in Hyperthermophilic Proteins----------13
1-7 Charged Amino Acids---------------------------------14
1-8 Electrostatic Interactions in RNA Recognition-------15
1-9 Post-Translational Modifications (PTMs)-------------18
1-10 Thesis Overview------------------------------------20
1-11 References-----------------------------------------22
Chapter 2.----------------------------------------------31
2-1 Introduction----------------------------------------31
β-Sheet-------------------------------------------------31
β-Hairpin-----------------------------------------------32
β-Turn--------------------------------------------------35
β-Sheet Propensity--------------------------------------37
Cross Strand Interaction--------------------------------39
Effect of Side Chain Length on Cross Strand Ion-pairs---41
Double Mutant Thermodynamic Cycle-----------------------42
2-2 Results and Discussion------------------------------43
Design of the Basic β-Hairpin Model---------------------43
Peptide Synthesis---------------------------------------45
Charcoal Mediated Air Oxidation of Fully Folded Peptides with Charcoal-------------------------------------------46
Sequence Specific Assignments---------------------------47
Secondary Structure Assignment by NMR Parameters--------53
Hairpin Stability---------------------------------------63
The Free Energy of Cross Strand Ion-pairs Interaction---66
2-3 Conclusions-----------------------------------------68
2-4 Acknowledgement-------------------------------------68
2-5 Experimental Section--------------------------------69
General Materials and Methods---------------------------69
Peptide Synthesis---------------------------------------70
Peptide Structure Analysis by 2D-NMR Spectrometry-------76
2-6 References------------------------------------------87
Chapter 3-----------------------------------------------93
3-1 Introduction----------------------------------------93
Ribonucleic acid (RNA)----------------------------------93
Human Immunodeficiency Virus (HIV)----------------------95
Trans-Activation Response Element (TAR) RNA-------------97
Trans-Activator of Transcription (Tat) Protein----------98
Tat-Activated Transcription----------------------------100
Inhibition of Tat-TAR Interaction----------------------101
Arginine Methylation in Tat-TAR Interaction------------103
Cell-penetrating Peptides------------------------------104
3-2 Results and Discussion-----------------------------107
Design of Tat-derived Peptides-------------------------107
Peptide Synthesis--------------------------------------109
Electrophoretic Mobility Shift Assay in the Presence of Bulk E. coli tRNA-------------------------------------------112
Tat-Derived Peptide Binding Affinity and Specificity towards TAR RNA------------------------------------------------119
Circular Dichroism Spectroscopy------------------------121
Cellular Uptake Assay----------------------------------121
3-3 Conclusion-----------------------------------------126
3-4 Acknowledgement------------------------------------126
3-5 Experimental Section-------------------------------127
General Materials and Methods--------------------------127
Peptide Synthesis--------------------------------------128
Ultraviolet-Visible (UV-vis) Spectroscopy--------------141
Gel Shift Assay----------------------------------------143
Circular Dichroism (CD) Spectroscopy-------------------145
Cells and Cell Cultures--------------------------------146
Cellular uptake Assay----------------------------------146
3-6 References-----------------------------------------148
Appendix-----------------------------------------------155

1-11 References
1.Crick, F. Central dogma of molecular biology. Nature 1970, 227, 561.
2.Invernizzi, G.; Papaleo, E.; Sabate, R.; Ventura, S. Protein aggregation: mechanisms and functional consequences. Int. J. Biochem. Cell. Biol. 2012, 44, 1541.
3.Ferri, C. P.; Prince, M.; Brayne, C.; Brodaty, H.; Fratiglioni, L.; Ganguli, M.; Hall, K.; Hasegawa, K.; Hendrie, H.; Huang, Y.; Jorm, A.; Mathers, C.; Menezes, P. R.; Rimmer, E.; Scazufca, M. Global prevalence of dementia: a Delphi consensus study. Lancet 2005, 366, 2112.
4.Prince, M.; Bryce, R.; Albanese, E.; Wimo, A.; Ribeiro, W.; Ferri, C. P. The global prevalence of dementia: A systematic review and metaanalysis. Alzheimers Dement. 2013, 9, 63.
5.Ballard, C.; Gauthier, S.; Corbett, A.; Brayne, C.; Aarsland, D.; Jones, E. Alzheimer''s disease. Lancet 2011, 377, 1019.
6.Nussbaum, R. L.; Ellis, C. E. Alzheimer''s disease and Parkinson''s disease. N. Engl. J. Med. 2003, 348, 1356.
7.Kahle, P. J. α-Synucleinopathy models and human neuropathology: similarities and differences. Acta. Neuropathol. 2008, 115, 87.
8.Caughey, B.; Chesebro, B. Prion protein and the transmissible spongiform encephalopathies. Trends. Cell. Biol. 1997, 7, 56.
9.Lee, S. J.; Lim, H. S.; Masliah, E.; Lee, H. J. Protein aggregate spreading in neurodegenerative diseases: problems and perspectives. Neurosci. Res. 2011, 70, 339.
10.Rao, S. T.; Rossmann, M. G. Comparison of super-secondary structures in proteins. J. Mol. Biol. 1973, 76, 241.
11.Bork, P.; Holm, L.; Sander, C. The immunoglobulin fold. Structural classification, sequence patterns and common core. J. Mol. Biol. 1994, 242, 309.
12.Wierenga, R. K. The TIM-barrel fold: a versatile framework for efficient enzymes. FEBS Lett. 2001, 492, 193.
13.Todd, A. E.; Orengo, C. A.; Thornton, J. M. Evolution of protein function, from a structural perspective. Opin. Chem. Biol. 1999, 3, 548.
14.Martin, A. C.; Orengo, C. A.; Hutchinson, E. G.; Jones, S.; Karmirantzou, M.; Laskowski, R. A.; Mitchell, J. B.; Taroni, C.; Thornton, J. M. Protein folds and functions. Structure 1998, 6, 875.
15.Linderstrom-Lang, K. U. Proteins and Enzymes. Lane. Medical. Lectures. 1952, 6.
16.Branden, C. T., J. Introduction to Protein Structure. In 2nd ed.; Garland Publishing: New York, 1999.
17.Pauling, L.; Corey, R. B. The pleated sheet, a new layer configuration of polypeptide chains. Proc. Natl. Acad. Sci. U. S. A. 1951, 37, 251.
18.Pauling, L.; Corey, R. B. The planarity of the amide group in polypeptides. J. Am. Chem. Soc. 1952, 74, 3964.
19.Ramachandran, G. N.; Ramakrishnan, C.; Sasisekharan, V. Stereochemistry of polypeptide chain configurations. J. Mol. Biol. 1963, 7, 95.
20.Richardson, J. S. The anatomy and taxonomy of protein structure. Adv. Protein. Chem. 1981, 34, 167.
21.Murzin, A. G.; Brenner, S. E.; Hubbard, T.; Chothia, C. SCOP: a structural classification of proteins database for the investigation of sequences and structures. J. Mol. Biol. 1995, 247, 536.
22.Pauling, L.; Corey, R. B.; Branson, H. R. The structure of proteins; two hydrogen-bonded helical configurations of the polypeptide chain. Proc. Natl. Acad. Sci. U. S. A. 1951, 37, 205.
23.Perutz, M. F. New x-ray evidence on the configuration of polypeptide chains. Nature 1951, 167, 1053.
24.Perutz, M. F.; Rossmann, M. G.; Cullis, A. F.; Muirhead, H.; Will, G.; North, A. C. Structure of haemoglobin: a three-dimensional fourier synthesis at 5.5-A. resolution, obtained by X-ray analysis. Nature 1960, 185, 416.
25.Arnott, S.; Wonacott, A. J. Atomic co-ordinates for an α-helix: refinement of the crystal structure of α-poly-l-alanine. J. Mol. Biol. 1966, 21, 371.
26.Barlow, D. J.; Thornton, J. M. Helix geometry in proteins. J. Mol. Biol. 1988, 201, 601.
27.Sibanda, B. L.; Thornton, J. M. β-hairpin families in globular proteins. Nature 1985, 316, 170.
28.Sibanda, B. L.; Blundell, T. L.; Thornton, J. M. Conformation of β-hairpins in protein structures. A systematic classification with applications to modelling by homology, electron density fitting and protein engineering. J. Mol. Biol. 1989, 206, 759.
29.Sibanda, B. L.; Thornton, J. M. Conformation of β-hairpins in protein structures: classification and diversity in homologous structures. Methods Enzymol. 1991, 202, 59.
30.Venkatachalam, C. M. Stereochemical criteria for polypeptides and proteins. V. Conformation of a system of three linked peptide units. Biopolymers 1968,6, 1425.
31.Lewis, P. N.; Momany, F. A.; Scheraga, H. A. Folding of polypeptide chains in proteins: a proposed mechanism for folding. Proc. Natl. Acad. Sci. U. S. A. 1971, 68, 2293.
32.Kuntz, I. D. Tertiary structure in carboxypeptidase. J. Am. Chem. Soc. 1972, 94, 8568.
33.Crawford, J. L.; Lipscomb, W. N.; Schellman, C. G. The reverse turn as a polypeptide conformation in globular proteins. Proc. Natl. Acad. Sci. U. S. A. 1973, 70, 538.
34.Levitt, M.; Chothia, C. Structural patterns in globular proteins. Nature 1976, 261, 552.
35.Chou, P. Y.; Fasman, G. D. β-Turns in Proteins. J. Mol. Biol. 1977, 115, 135.
36.Rose, G. D.; Seltzer, J. P. A new algorithm for finding the peptide chain turns in a globular protein. J. Mol. Biol. 1977, 113, 153.
37.Lewis, P. N.; Momany, F. A.; Scheraga, H. A. Chain reversals in proteins. Biochim. Biophys. Acta. 1973, 303, 211.
38.Chou, P. Y.; Fasman, G. D. Prediction of protein conformation. Biochemistry 1974, 13, 222.
39.Janin, J.; Chothia, C. Domains in proteins: definitions, location, and structural principles. Methods Enzymol. 1985, 115, 420.
40.Phillips, D. C. The three-dimensional structure of an enzyme molecule. Sci. Am. 1966, 215, 78.
41.Drenth, J.; Jansonius, J. N.; Koekoek, R.; Swen, H. M.; Wolthers, B. G. Structure of papain. Nature 1968, 218, 929.
42.Davies, D. R.; Padlan, E. A.; Segal, D. M. Three-dimensional structure of immunoglobulins. Annu. Rev. Biochem. 1975, 44, 639.
43.Amzel, L. M.; Poljak, R. J. Three-dimensional structure of immunoglobulins. Annu. Rev. Biochem. 1979, 48, 961.
44.Torres, M.; Casadevall, A. The immunoglobulin constant region contributes to affinity and specificity. Trends Immunol. 2008, 29, 91.
45.Klotz, I. M.; Langerman, N. R.; Darnall, D. W. Quaternary structure of proteins. Annu. Rev. Biochem. 1970, 39, 25.
46.Baldwin, R. L.; Rose, G. D. Is protein folding hierarchic? I. Local structure and peptide folding. Trends. Biochem. Sci. 1999, 24, 26.
47.Baldwin, R. L.; Rose, G. D. Is protein folding hierarchic? II. Folding intermediates and transition states. Trends. Biochem. Sci. 1999, 24, 77.
48.Cantor, C. R. S., P. R. Biophysical Chemistry (Part I-III) Part I: The conformation of biological macromolecules. Part II: Techniques for the study of biological structure and function. Part III: The behavior of biological macromolecules. In Freeman, W. H. and Co.: New York, 1980.
49.Creighton, T. E. Proteins: Structures and molecular properties. In Second ed.; Freeman, W. H.: New York, 1993.
50.Dill, K. A. Dominant forces in protein folding. Biochemistry 1990, 29, 7133.
51.Hagler, A. T.; Huler, E.; Lifson, S. Energy functions for peptides and proteins. I. Derivation of a consistent force field including the hydrogen bond from amide crystals. J. Am. Chem. Soc. 1974, 96, 5319.
52.Perutz, M. F. Electrostatic effects in proteins. Science 1978, 201, 1187.
53.Barlow, D. J.; Thornton, J. M. Ion-pairs in proteins. J. Mol. Biol. 1983, 168, 867.
54.Nicholls, A.; Sharp, K. A.; Honig, B. Protein folding and association: insights from the interfacial and thermodynamic properties of hydrocarbons. Proteins 1991, 11, 281.
55.Stickle, D. F.; Presta, L. G.; Dill, K. A.; Rose, G. D. Hydrogen bonding in globular proteins. J. Mol. Biol. 1992, 226, 1143.
56.Pace, C. N.; Shirley, B. A.; McNutt, M.; Gajiwala, K. Forces contributing to the conformational stability of proteins. FASEB J. 1996, 10, 75.
57.Pace, C. N.; Grimsley, G. R.; Scholtz, J. M. Protein ionizable groups: pK values and their contribution to protein stability and solubility. J. Biol. Chem. 2009, 284, 13285.
58.Stigter, D.; Dill, K. A. Charge effects on folded and unfolded proteins. Biochemistry 1990, 29, 1262.
59.Yoder, C. H. Teaching ion-ion, ion-dipole, and dipole-dipole interactions. J. Chem. Educ. 1977, 54, 402.
60.Wada, A.; Nakamura, H. Nature of the charge distribution in proteins. Nature 1981, 293, 757.
61.Hol, W. G.; Halie, L. M.; Sander, C. Dipoles of the α-helix and β-sheet: their role in protein folding. Nature 1981, 294, 532.
62.Hol, W. G.; van Duijnen, P. T.; Berendsen, H. J. The α-helix dipole and the properties of proteins. Nature 1978, 273, 443.
63.van Duijnen, P. T.; Thole, B. T.; Hol, W. G. On the role of the active site helix in papain, an ab initio molecular orbital study. Biophys. Chem. 1979, 9, 273.
64.Sheridan, R. P.; Allen, L. C. The electrostatic potential of the α-helix (electrostatic potential/α-helix/secondary structure/helix dipole). Biophys. Chem. 1980, 11, 133.
65.Atkins, P. W. J., L. Chemical principles: the quest for insight. In fourth ed.; Freeman W.H. : New York, 2008.
66.Hagler, A. T.; Lifson, S. Energy functions for peptides and proteins. II. The amide hydrogen bond and calculation of amide crystal properties. J. Am. Chem. Soc. 1974, 96, 5327.
67.Baker, E. N.; Hubbard, R. E. Hydrogen bonding in globular proteins. Prog. Biophys. Mol. Biol. 1984, 44, 97.
68.McDonald, I. K.; Thornton, J. M. Satisfying hydrogen bonding potential in proteins. J. Mol. Biol. 1994, 238, 777.
69.Klapper, M. H. On the nature of the protein interior. Biochim. Biophys. Acta. 1971, 229, 557.
70.Richards, F. M. Areas, volumes, packing and protein structure. Annu. Rev. Biophys. Bioeng. 1977, 6, 151.
71.Lesser, G. J.; Rose, G. D. Hydrophobicity of amino acid subgroups in proteins. Proteins 1990, 8, 6.
72.Harpaz, Y.; Gerstein, M.; Chothia, C. Volume changes on protein folding. Structure 1994, 2, 641.
73.Kauzmann, W. Some factors in the interpretation of protein denaturation. Adv. Protein Chem. 1959, 14, 1.
74.Tanford, C. Contribution of hydrophobic interactions to the stability of the globular conformation of proteins. J. Am. Chem. Soc. 1962, 84, 4240.
75.Pace, C. N. Contribution of the hydrophobic effect to globular protein stability. J. Mol. Biol. 1992, 226, 29.
76.Fersht, A. R. S. Principles of protein stability derived from protein engineering experiments. . Curr. Opin. Struct. Biol. 1993, 3, 75.
77.Honig, B.; Yang, A. S. Free energy balance in protein folding. Adv. Protein Chem. 1995, 46, 27.
78.Lazaridis, T.; Archontis, G.; Karplus, M. Enthalpic contribution to protein stability: insights from atom-based calculations and statistical mechanics. Adv. Protein Chem. 1995, 47, 231.
79.Makhatadze, G. I.; Privalov, P. L. Energetics of protein structure. Adv. Protein. Chem. 1995, 47, 307.
80.Matthews, B. W. Studies on protein stability with T4 lysozyme. Adv. Protein Chem. 1995, 46, 249.
81.Feinberg, G.; Sucher, J. General theory of the van der Waals interaction: A model-independent approach. Phys. Rev. A Gen. Phys. 1970, 2, 2395.
82.Levitt, M.; Gerstein, M.; Huang, E.; Subbiah, S.; Tsai, J. Protein folding: the endgame. Annu. Rev. Biochem. 1997, 66, 549.
83.Amelunxen, R. E. Some chemical and physical properties of thermostable glyceraldehyde-3-phosphate dehydrogenase from Bacillus stearothermophilus. Biochim. Biophys. Acta. 1967, 139, 24.
84.Brock, T. D. Life at high temperatures. Evolutionary, ecological, and biochemical significance of organisms living in hot springs is discussed. Science 1967, 158, 1012.
85.Suzuki, K.; Ieuan Harris, J. Glyceraldehyde-3-phosphate dehydrogenase from Bacillus stearothermophilus. FEBS Lett. 1971, 13, 217.
86.Valentine, R. C. Bacterial ferredoxin. Bacteriol. Rev. 1964, 28, 497.
87.Devanath.T; Akagi, J. M.; Hersh, R. T.; Himes, R. H. Ferredoxin from 2 thermophilic clostridia. J. Biol. Chem. 1969, 244, 2846.
88.Kunkel, H. G.; Wallenius, G. New hemoglobin in normal adult blood. Science 1955, 122, 288.
89.Kinderle.J; Lehmann, H.; Tipton, K. F. Thermal denaturation of human oxyhemoglobins a, A2, C and S. Biochem. J. 1973, 135, 805.
90.Efstratiadis, A.; Posakony, J. W.; Maniatis, T.; Lawn, R. M.; Oconnell, C.; Spritz, R. A.; Deriel, J. K.; Forget, B. G.; Weissman, S. M.; Slightom, J. L.; Blechl, A. E.; Smithies, O.; Baralle, F. E.; Shoulders, C. C.; Proudfoot, N. J. The structure and evolution of the human β-Globin gene family. Cell 1980, 21, 653.
91.Perutz, M. F.; Raidt, H. Stereochemical basis of heat stability in bacterial ferredoxins and in haemoglobin A2. Nature 1975, 255, 256.
92.Yip, K. S.; Stillman, T. J.; Britton, K. L.; Artymiuk, P. J.; Baker, P. J.; Sedelnikova, S. E.; Engel, P. C.; Pasquo, A.; Chiaraluce, R.; Consalvi, V.; et al. The structure of Pyrococcus furiosus glutamate dehydrogenase reveals a key role for ion-pair networks in maintaining enzyme stability at extreme temperatures. Structure 1995, 3, 1147.
93.Jaenicke, R.; Schurig, H.; Beaucamp, N.; Ostendorp, R. Structure and stability of hyperstable proteins: glycolytic enzymes from hyperthermophilic bacterium Thermotoga maritima. Adv. Protein. Chem. 1996, 48, 181.
94.Hendsch, Z. S.; Tidor, B. Do salt bridges stabilize proteins? A continuum electrostatic analysis. Protein Sci. 1994, 3, 211.
95.Blake, C. C.; Mair, G. A.; North, A. C.; Phillips, D. C.; Sarma, V. R. On the conformation of the hen egg-white lysozyme molecule. Proc. R. Soc. Lond. B. Biol. Sci. 1967, 167, 365.
96.Diamond, R. Real-space refinement of the structure of hen egg-white lysozyme. J. Mol. Biol. 1974, 82, 371.
97.Mauguen, Y.; Hartley, R. W.; Dodson, E. J.; Dodson, G. G.; Bricogne, G.; Chothia, C.; Jack, A. Molecular structure of a new family of ribonucleases. Nature 1982, 297, 162.
98.Weaver, L. H.; Matthews, B. W. Structure of bacteriophage T4 lysozyme refined at 1.7 A resolution. J. Mol. Biol. 1987, 193, 189.
99.Morize, I.; Surcouf, E.; Vaney, M. C.; Epelboin, Y.; Buehner, M.; Fridlansky, F.; Milgrom, E.; Mornon, J. P. Refinement of the C222(1) crystal form of oxidized uteroglobin at 1.34 A resolution. J. Mol. Biol. 1987, 194, 725.
100.Vijaykumar, S.; Bugg, C. E.; Cook, W. J. Structure of ubiquitin refined at 1.8 a resolution. J. Mol. Biol. 1987, 194, 531.
101.Corfield, P. W.; Lee, T. J.; Low, B. W. The crystal structure of erabutoxin a at 2.0 A resolution. J. Biol. Chem. 1989, 264, 9239.
102.Mondragon, A.; Subbiah, S.; Almo, S. C.; Drottar, M.; Harrison, S. C. Structure of the amino-terminal domain of phage 434 repressor at 2.0 A resolution. J. Mol. Biol. 1989, 205, 189.
103.O''Shea, E. K.; Klemm, J. D.; Kim, P. S.; Alber, T. X-ray structure of the GCN4 leucine zipper, a two-stranded, parallel coiled coil. Science 1991, 254, 539.
104.Beamer, L. J.; Pabo, C. O. Refined 1.8 A crystal structure of the lambda repressor-operator complex. J. Mol. Biol. 1992, 227, 177.
105.Matsuo, H.; Li, H.; McGuire, A. M.; Fletcher, C. M.; Gingras, A. C.; Sonenberg, N.; Wagner, G. Structure of translation factor eIF4E bound to m7GDP and interaction with 4E-binding protein. Nat. Struct. Biol. 1997, 4, 717.
106.Varani, G.; Nagai, K. RNA recognition by RNP proteins during RNA processing. Annu. Rev. Biophys. Biomol. Struct. 1998, 27, 407.
107.Seeman, N. C.; Rosenberg, J. M.; Rich, A. Sequence-specific recognition of double helical nucleic acids by proteins. Proc. Natl. Acad. Sci. U. S. A. 1976, 73, 804.
108.Dickerson, R. E.; Drew, H. R.; Conner, B. N.; Wing, R. M.; Fratini, A. V.; Kopka, M. L. The anatomy of A-, B-, and Z-DNA. Science 1982, 216, 475.
109.Seeman, N. C.; Rosenberg, J. M.; Rich, A. Sequence-specific recognition of double helical nucleic acids by proteins. Proc. Natl. Acad. Sci. U. S. A. 1976, 73, 804.
110.Schleif, R. DNA binding by proteins. Science 1988, 241, 1182.
111.Stefl, R.; Allain, F. H. A novel RNA pentaloop fold involved in targeting ADAR2. RNA 2005, 11, 592.
112.Drew, H. R.; Wing, R. M.; Takano, T.; Broka, C.; Tanaka, S.; Itakura, K.; Dickerson, R. E. Structure of a B-DNA dodecamer: conformation and dynamics. Proc. Natl. Acad. Sci. U. S. A. 1981, 78, 2179.
113.Varmus, H. Retroviruses. Science 1988, 240, 1427.
114.Weiss, R. A. How does HIV cause AIDS. Science 1993, 260, 1273.
115.Greene, W. C. The molecular biology of human immunodeficiency virus type 1 infection. N. Engl. J. Med. 1991,324, 308.
116.Weeks, K. M.; Ampe, C.; Schultz, S. C.; Steitz, T. A.; Crothers, D. M. Fragments of the HIV-1 Tat protein specifically bind TAR RNA. Science 1990, 249, 1281.
117.Berkhout, B.; Jeang, K. T. trans activation of human immunodeficiency virus type 1 is sequence specific for both the single-stranded bulge and loop of the trans-acting-responsive hairpin: a quantitative analysis. J. Virol. 1989, 63, 5501.
118.Merrick, W. C. Mechanism and regulation of eukaryotic protein synthesis. Microbiol Rev 1992, 56, 291.
119.Uy, R.; Wold, F. Post-translational covalent modification of proteins. Science 1977, 198, 890.
120.Seo, J.; Lee, K. J. Post-translational modifications and their biological functions: proteomic analysis and systematic approaches. J. Biochem. Mol. Biol. 2004, 37, 35.
121.Desrosiers, R.; Tanguay, R. M. Methylation of Drosophila histones at proline, lysine, and arginine residues during heat shock. J Biol Chem 1988, 263, 4686.
122.Najbauer, J.; Orpiszewski, J.; Aswad, D. W. Molecular aging of tubulin: accumulation of isoaspartyl sites in vitro and in vivo. Biochemistry 1996, 35, 5183.
123.Chen, D.; Ma, H.; Hong, H.; Koh, S. S.; Huang, S. M.; Schurter, B. T.; Aswad, D. W.; Stallcup, M. R. Regulation of transcription by a protein methyltransferase. Science 1999, 284, 2174.
124.Paik, W. K.; Kim, S. Protein methylation. Science 1971, 174, 114.

2-6 References
1.Ramachandran, G. N.; Ramakrishnan, C.; Sasisekharan, V. Stereochemistry of polypeptide chain configurations. J. Mol. Biol. 1963, 7, 95.
2.Pauling, L.; Corey, R. B. The pleated sheet, a new layer configuration of polypeptide chains. Proc. Natl. Acad. Sci. U. S. A. 1951, 37, 251.
3.Richardson, J. S. β-Sheet topology and the relatedness of proteins. Nature 1977, 268, 495.
4.Sibanda, B. L.; Thornton, J. M. β-Hairpin families in globular proteins. Nature 1985, 316, 170.
5.Sibanda, B. L.; Blundell, T. L.; Thornton, J. M. Conformation of β-hairpins in protein structures. A systematic classification with applications to modelling by homology, electron density fitting and protein engineering. J. Mol. Biol. 1989, 206, 759.
6.Butterfield, S. M.; Waters, M. L. A designed β-hairpin peptide for molecular recognition of ATP in water. J. Am. Chem. Soc. 2003, 125, 9580.
7.Minor, D. L., Jr.; Kim, P. S. Measurement of the β-sheet-forming propensities of amino acids. Nature 1994, 367, 660.
8.Minor, D. L., Jr.; Kim, P. S. Context is a major determinant of β-sheet propensity. Nature 1994, 371, 264.
9.Smith, C. K.; Withka, J. M.; Regan, L. A thermodynamic scale for the β-sheet forming tendencies of the amino acids. Biochemistry 1994, 33, 5510.
10.Gellman, S. H. Minimal model systems for β-sheet secondary structure in proteins. Curr. Opin. Chem. Biol. 1998, 2, 717.
11.Blanco, F. J. J., M. A.; Herranz, J.; Roco, M.; Santoro, J.; Nieto, J. L. NMR evidence of a short linear peptide that folds into a β-hairpin in aqueous solution. J. Am. Chem. Soc. 1993, 115, 5887.
12.Blanco, F. J.; Rivas, G.; Serrano, L. A short linear peptide that folds into a native stable β-hairpin in aqueous solution. Nat. Struct. Biol. 1994, 1, 584.
13.Searle, M. S.; Williams, D. H.; Packman, L. C. A short linear peptide derived from the N-terminal sequence of ubiquitin folds into a water-stable non-native β-hairpin. Nat. Struct. Biol. 1995, 2, 999.
14.Venkatachalam, C. M. Stereochemical criteria for polypeptides and proteins. V. Conformation of a system of three linked peptide units. Biopolymers 1968, 6, 1425.
15.Crawford, J. L.; Lipscomb, W. N.; Schellman, C. G. The reverse turn as a polypeptide conformation in globular proteins. Proc. Natl. Acad. Sci. U. S. A. 1973, 70, 538.
16.Chou, P. Y.; Fasman, G. D. β-Turns in proteins. J. Mol. Biol. 1977, 115, 135.
17.Kabsch, W.; Sander, C. Dictionary of protein secondary structure: pattern recognition of hydrogen-bonded and geometrical features. Biopolymers 1983, 22, 2577.
18.Lewis, P. N.; Momany, F. A.; Scheraga, H. A. Chain reversals in proteins. Biochim. Biophys. Acta. 1973, 303, 211.
19.Chou, P. Y.; Fasman, G. D. Prediction of protein conformation. Biochemistry 1974, 13, 222.
20.Rose, G. D.; Gierasch, L. M.; Smith, J. A. Turns in peptides and proteins. Adv. Protein Chem. 1985, 37, 1.
21.IUPAC-IUB Commission on biochemical nomenclature. Abbreviations and symbols for the description of the conformation of polypeptide chains. Tentative rules (1969). Biochemistry 1970, 9, 3471.
22.Richardson, J. S. The anatomy and taxonomy of protein structure. Adv. Protein Chem. 1981, 34, 167.
23.Hutchinson, E. G.; Thornton, J. M. A revised set of potentials for β-turn formation in proteins. Protein Sci. 1994, 3, 2207.
24.Griffiths-Jones, S. R.; Maynard, A. J.; Searle, M. S. Dissecting the stability of a β-hairpin peptide that folds in water: NMR and molecular dynamics analysis of the β-turn and β-strand contributions to folding. J. Mol. Biol. 1999, 292, 1051.
25.Stanger, H. E. G., S. H. Rules for antiparallel β-sheet design: D-Pro-Gly is superior to L-Asn-Gly for β-hairpin nucleation. J. Am. Chem. Soc. 1998, 120, 4236.
26.Chou, P. Y.; Fasman, G. D. Conformational parameters for amino acids in helical, β-sheet, and random coil regions calculated from proteins. Biochemistry 1974, 13, 211.
27.Kim, C. A.; Berg, J. M. Thermodynamic β-sheet propensities measured using a zinc-finger host peptide. Nature 1993, 362, 267.
28.Ramirez-Alvarado, M.; Kortemme, T.; Blanco, F. J.; Serrano, L. β-Hairpin and β-sheet formation in designed linear peptides. Bioorg. Med. Chem. 1999, 7, 93.
29.Wouters, M. A.; Curmi, P. M. An analysis of side chain interactions and pair correlations within antiparallel β-sheets: the differences between backbone hydrogen-bonded and non-hydrogen-bonded residue pairs. Proteins 1995, 22, 119.
30.Syud, F. A.; Stanger, H. E.; Gellman, S. H. Interstrand side chain-side chain interactions in a designed β-hairpin: significance of both lateral and diagonal pairings. J. Am. Chem. Soc. 2001, 123, 8667.
31.Dill, K. A. Dominant forces in protein folding. Biochemistry 1990, 29, 7133.
32.Searle, M. S.; Griffiths-Jones, S. R.; Skinner-Smith, H. Energetics of weak interactions in a β-hairpin peptide: Electrostatic and hydrophobic contributions to stability from lysine salt bridges. J. Am. Chem. Soc. 1999, 121, 11615.
33.Ciani, B.; Jourdan, M.; Searle, M. S. Stabilization of β-hairpin peptides by salt bridges: role of preorganization in the energetic contribution of weak interactions. J. Am. Chem. Soc. 2003, 125, 9038.
34.Deechongkit, S.; Nguyen, H.; Powers, E. T.; Dawson, P. E.; Gruebele, M.; Kelly, J. W. Context-dependent contributions of backbone hydrogen bonding to β-sheet folding energetics. Nature 2004, 430, 101.
35.Tatko, C. D.; Waters, M. L. Comparison of C-H...Π and hydrophobic interactions in a β-hairpin peptide: impact on stability and specificity. J. Am. Chem. Soc. 2004, 126, 2028.
36.Kiehna, S. E.; Waters, M. L. Sequence dependence of β-hairpin structure: comparison of a salt bridge and an aromatic interaction. Protein Sci. 2003, 12, 2657.
37.Barlow, D. J.; Thornton, J. M. Ion-pairs in proteins. J. Mol. Biol. 1983, 168, 867.
38.Padmanabhan, S.; York, E. J.; Stewart, J. M.; Baldwin, R. L. Helix propensities of basic amino acids increase with the length of the side-chain. J. Mol. Biol. 1996, 257, 726.
39.Cheng, R. P.; Weng, Y. J.; Wang, W. R.; Koyack, M. J.; Suzuki, Y.; Wu, C. H.; Yang, P. A.; Hsu, H. C.; Kuo, H. T.; Girinath, P.; Fang, C. J. Helix formation and capping energetics of arginine analogs with varying side chain length. Amino Acids 2012, 43, 195.
40.Hughes, R. M.; Benshoff, M. L.; Waters, M. L. Effects of chain length and N-methylation on a cation-Π interaction in a β-hairpin peptide. Chemistry 2007, 13, 5753.
41.Chakrabartty, A.; Kortemme, T.; Baldwin, R. L. Helix propensities of the amino acids measured in alanine-based peptides without helix-stabilizing side-chain interactions. Protein Sci. 1994, 3, 843.
42.Syud, F. A.; Espinosa, J. F.; Gellman, S. H. NMR-based quantification of β-sheet populations in aqueous solution through use of reference peptides for the folded and unfolded states. J. Am. Chem. Soc. 1999, 121, 11577.
43.Carter, P. J.; Winter, G.; Wilkinson, A. J.; Fersht, A. R. The use of double mutants to detect structural changes in the active site of the tyrosyl-tRNA synthetase (Bacillus stearothermophilus). Cell 1984, 38, 835.
44.Ackers, G. K.; Smith, F. R. Effects of site-specific amino acid modification on protein interactions and biological function. Annu. Rev. Biochem. 1985, 54, 597.
45.Horovitz, A.; Fersht, A. R. Strategy for analysing the co-operativity of intramolecular interactions in peptides and proteins. J. Mol. Biol. 1990, 214, 613.
46.Serrano, L.; Horovitz, A.; Avron, B.; Bycroft, M.; Fersht, A. R. Estimating the contribution of engineered surface electrostatic interactions to protein stability by using double-mutant cycles. Biochemistry 1990, 29, 9343.
47.Atherton, E.; Fox, H.; Harkiss, D.; Logan, C. J.; Sheppard, R. C.; Williams, B. J. A mild procedure for solid phase peptide synthesis: use of fluorenylmethoxycarbonylamino-acids. J. Chem. Soc., Chem. Commun. 1978, 537.
48.Fields, G. B.; Noble, R. L. Solid phase peptide synthesis utilizing 9-fluorenylmethoxycarbonyl amino acids. Int. J. Pept. Protein Res. 1990,35, 161.
49.Volkmer-Engert, R.; Landgraf, C.; Schneider-Mergener, J. Charcoal surface-assisted catalysis of intramolecular disulfide bond formation in peptides. J. Pept. Res. 1998, 51, 365.
50.Bax, A. D., D. G. . MLEV-17-based two-dimensional homonuclear magnetization transfer spectroscopy. J. Magn. Reson. 1985, 65.
51.Aue, W. P. B., E.; Ernst, R. R. Two-dimensional spectroscopy- application to nuclear magnetic-resonance. J. Chem. Phys. 1976, 64, 2229.
52.Feichtinger, K. Z., C.; Sings, H. L..; Goodman, M. Diprotected triflylguanidines: A new class of guanidinylation reagents. J. Org. Chem. 1998,63, 3804.
53.Bundi, A. W., K. 1H-nmr parameters of the common amino acid residues measured in aqueous solutions of the linear tetrapeptides H-Gly-Gly-X-L-Ala-OH. Biopolymers 1979, 18, 285.
54.Yao, J.; Dyson, H. J.; Wright, P. E. Chemical shift dispersion and secondary structure prediction in unfolded and partly folded proteins. FEBS Lett. 1997, 419, 285.
55.Markley, J. L.; Meadows, D. H.; Jardetzky, O. Nuclear magnetic resonance studies of helix-coil transitions in polyamino acids. J. Mol. Biol. 1967, 27, 25.
56.Nakamura, A.; Jardetzky, O. Systematic analysis of chemical shifts in the nuclear magnetic resonance spectra of peptide chains. II. Oligoglycines. Biochemistry 1968, 7, 1226.
57.Wishart, D. S.; Sykes, B. D.; Richards, F. M. Relationship between nuclear magnetic resonance chemical shift and protein secondary structure. J. Mol. Biol. 1991, 222, 311.
58.Wishart, D. S.; Sykes, B. D.; Richards, F. M. The chemical shift index: a fast and simple method for the assignment of protein secondary structure through NMR spectroscopy. Biochemistry 1992, 31, 1647.
59.Wuthrich, K. NMR of proteins and nucleic acids. 1986.
60.Lukin, J. A.; Gove, A. P.; Talukdar, S. N.; Ho, C. Automated probabilistic method for assigning backbone resonances of (13C, 15N)-labeled proteins. J. Biomol. NMR 1997, 9, 151.
61.Schwarzinger, S.; Kroon, G. J.; Foss, T. R.; Wright, P. E.; Dyson, H. J. Random coil chemical shifts in acidic 8 M urea: implementation of random coil shift data in NMR View. J. Biomol. NMR 2000, 18, 43.
62.Delepierre, M.; Dobson, C. M.; Poulsen, F. M. Studies of β-sheet structure in lysozyme by proton nuclear magnetic resonance. Assignments and analysis of spin-spin coupling constants. Biochemistry 1982, 21, 4756.
63.Wishart, D. S.; Bigam, C. G.; Holm, A.; Hodges, R. S.; Sykes, B. D. (1)H, (13)C and (15)N random coil NMR chemical shifts of the common amino acids. I. Investigations of nearest-neighbor effects. J. Biomol. NMR 1995, 5, 332.
64.Kim, Y. M.; Prestegard, J. H. Measurement of vicinal couplings from cross peaks in cosy spectra. J. Magn. Reson. 1989, 84, 9.
65.Jeener, J. M., B. H.; Bachmann, P.; Ernst, R. R. Investigation of exchange processes by two-dimensional NMR spectroscopy. J. Chem. Phys. 1979, 71, 4546.
3-6 References
1.Fresco, J. R.; Alberts, B. M.; Doty, P. Some molecular details of the secondary structure of ribonucleic acid. Nature 1960, 188, 98.
2.Ding, Y.; Lawrence, C. E. A statistical sampling algorithm for RNA secondary structure prediction. Nucleic Acids Res. 2003, 31, 7280.
3.Central dogma reversed. Nature 1970, 226, 1198.
4.Hu, W. S.; Temin, H. M. Retroviral recombination and reverse transcription. Science 1990, 250, 1227.
5.Varmus, H. Retroviruses. Science 1988, 240, 1427.
6.Weiss, R. A. How does HIV cause AIDS. Science 1993, 260, 1273.
7.Greene, W. C. The molecular biology of human immunodeficiency virus type 1 infection. N. Engl. J. Med. 1991, 324, 308.
8.Greene, W. C.; Peterlin, B. M. Charting HIV''s remarkable voyage through the cell: Basic science as a passport to future therapy. Nat. Med. 2002, 8, 673.
9.Chan, D. C.; Kim, P. S. HIV entry and its inhibition. Cell 1998, 93, 681.
10.Miller, M. D.; Farnet, C. M.; Bushman, F. D. Human immunodeficiency virus type 1 preintegration complexes: studies of organization and composition. J. Virol. 1997, 71, 5382.
11.Frankel, A. D.; Young, J. A. HIV-1: fifteen proteins and an RNA. Annu. Rev. Biochem. 1998, 67, 1.
12.Stoff, D. M.; Khalsa, J. H.; Monjan, A.; Portegies, P. Introduction: HIV/AIDS and Aging. AIDS 2004, 18 Suppl 1, S1.
13.Draper, D. E. Protein-RNA recognition. Annu. Rev. Biochem. 1995, 64, 593.
14.De Guzman, R. N.; Turner, R. B.; Summers, M. F. Protein-RNA recognition. Biopolymers 1998, 48, 181.
15.Fisher, A. G.; Feinberg, M. B.; Josephs, S. F.; Harper, M. E.; Marselle, L. M.; Reyes, G.; Gonda, M. A.; Aldovini, A.; Debouk, C.; Gallo, R. C.; et al. The trans-activator gene of HTLV-III is essential for virus replication. Nature 1986, 320, 367.
16.Cullen, B. R.; Greene, W. C. Regulatory pathways governing HIV-1 replication. Cell 1989, 58, 423.
17.Steffy, K.; Wong-Staal, F. Genetic regulation of human immunodeficiency virus. Microbiol. Rev. 1991, 55, 193.
18.Cullen, B. R. Regulation of HIV-1 gene expression. FASEB J. 1991, 5, 2361.
19.Rosen, C. A.; Sodroski, J. G.; Haseltine, W. A. The location of cis-acting regulatory sequences in the human T cell lymphotropic virus type III (HTLV-III/LAV) long terminal repeat. Cell 1985, 41, 813.
20.Sodroski, J.; Patarca, R.; Rosen, C.; Wong-Staal, F.; Haseltine, W. Location of the trans-activating region on the genome of human T-cell lymphotropic virus type III. Science 1985, 229, 74.
21.Arya, S. K.; Guo, C.; Josephs, S. F.; Wong-Staal, F. Trans-activator gene of human T-lymphotropic virus type III (HTLV-III). Science 1985, 229, 69.
22.Sodroski, J.; Rosen, C.; Wong-Staal, F.; Salahuddin, S. Z.; Popovic, M.; Arya, S.; Gallo, R. C.; Haseltine, W. A. Trans-acting transcriptional regulation of human T-cell leukemia virus type III long terminal repeat. Science 1985, 227, 171.
23.Selby, M. J.; Bain, E. S.; Luciw, P. A.; Peterlin, B. M. Structure, sequence, and position of the stem-loop in tar determine transcriptional elongation by tat through the HIV-1 long terminal repeat. Genes Dev. 1989, 3, 547.
24.Muesing, M. A.; Smith, D. H.; Capon, D. J. Regulation of mRNA accumulation by a human immunodeficiency virus trans-activator protein. Cell 1987, 48, 691.
25.Peterlin, B. M.; Luciw, P. A.; Barr, P. J.; Walker, M. D. Elevated levels of mRNA can account for the trans-activation of human immunodeficiency virus. Proc. Natl. Acad. Sci. U. S. A. 1986, 83, 9734.
26.Cordingley, M. G.; LaFemina, R. L.; Callahan, P. L.; Condra, J. H.; Sardana, V. V.; Graham, D. J.; Nguyen, T. M.; LeGrow, K.; Gotlib, L.; Schlabach, A. J.; et al. Sequence-specific interaction of Tat protein and Tat peptides with the transactivation-responsive sequence element of human immunodeficiency virus type 1 in vitro. Proc. Natl. Acad. Sci. U. S. A. 1990, 87, 8985.
27.Dingwall, C.; Ernberg, I.; Gait, M. J.; Green, S. M.; Heaphy, S.; Karn, J.; Lowe, A. D.; Singh, M.; Skinner, M. A. HIV-1 tat protein stimulates transcription by binding to a U-rich bulge in the stem of the TAR RNA structure. EMBO J. 1990, 9, 4145.
28.Roy, S.; Delling, U.; Chen, C. H.; Rosen, C. A.; Sonenberg, N. A bulge structure in HIV-1 TAR RNA is required for Tat binding and Tat-mediated trans-activation. Genes Dev. 1990, 4, 1365.
29.Weeks, K. M.; Ampe, C.; Schultz, S. C.; Steitz, T. A.; Crothers, D. M. Fragments of the HIV-1 Tat protein specifically bind TAR RNA. Science 1990, 249, 1281.
30.Muller, W. E.; Okamoto, T.; Reuter, P.; Ugarkovic, D.; Schroder, H. C. Functional characterization of Tat protein from human immunodeficiency virus. Evidence that Tat links viral RNAs to nuclear matrix. J. Biol. Chem. 1990, 265, 3803.
31.Gatignol, A.; Kumar, A.; Rabson, A.; Jeang, K. T. Identification of cellular proteins that bind to the human immunodeficiency virus type 1 trans-activation-responsive TAR element RNA. Proc. Natl. Acad. Sci. U. S. A. 1989, 86, 7828.
32.Marciniak, R. A.; Garcia-Blanco, M. A.; Sharp, P. A. Identification and characterization of a HeLa nuclear protein that specifically binds to the trans-activation-response (TAR) element of human immunodeficiency virus. Proc. Natl. Acad. Sci. U. S. A. 1990, 87, 3624.
33.Gaynor, R.; Soultanakis, E.; Kuwabara, M.; Garcia, J.; Sigman, D. S. Specific binding of a HeLa cell nuclear protein to RNA sequences in the human immunodeficiency virus transactivating region. Proc. Natl. Acad. Sci. U. S. A. 1989, 86, 4858.
34.Sheline, C. T.; Milocco, L. H.; Jones, K. A. Two distinct nuclear transcription factors recognize loop and bulge residues of the HIV-1 TAR RNA hairpin. Genes Dev. 1991, 5, 2508.
35.Wu, F.; Garcia, J.; Sigman, D.; Gaynor, R. tat regulates binding of the human immunodeficiency virus trans-activating region RNA loop-binding protein TRP-185. Genes Dev. 1991, 5, 2128.
36.Weeks, K. M.; Crothers, D. M. RNA recognition by Tat-derived peptides: interaction in the major groove? Cell 1991, 66, 577.
37.Churcher, M. J.; Lamont, C.; Hamy, F.; Dingwall, C.; Green, S. M.; Lowe, A. D.; Butler, J. G.; Gait, M. J.; Karn, J. High affinity binding of TAR RNA by the human immunodeficiency virus type-1 tat protein requires base-pairs in the RNA stem and amino acid residues flanking the basic region. J. Mol. Biol. 1993, 230, 90.
38.Jeang, K. T.; Gatignol, A. Comparison of regulatory features among primate lentiviruses. Curr. Top. Microbiol. Immunol. 1994, 188, 123.
39.Rana, T. M.; Jeang, K. T. Biochemical and functional interactions between HIV-1 Tat protein and TAR RNA. Arch. Biochem. Biophys. 1999, 365, 175.
40.Frankel, A. D.; Biancalana, S.; Hudson, D. Activity of synthetic peptides from the Tat protein of human immunodeficiency virus type 1. Proc. Natl. Acad. Sci. U. S. A. 1989, 86, 7397.
41.Kuppuswamy, M.; Subramanian, T.; Srinivasan, A.; Chinnadurai, G. Multiple functional domains of Tat, the trans-activator of HIV-1, defined by mutational analysis. Nucleic Acids Res. 1989, 17, 3551.
42.Hauber, J.; Perkins, A.; Heimer, E. P.; Cullen, B. R. Trans-activation of human immunodeficiency virus gene expression is mediated by nuclear events. Proc. Natl. Acad. Sci. U. S. A. 1987, 84, 6364.
43.Dang, C. V.; Lee, W. M. Nuclear and nucleolar targeting sequences of c-erb-A, c-myb, N-myc, p53, HSP70, and HIV tat proteins. J. Biol. Chem. 1989, 264, 18019.
44.Endo, S.; Kubota, S.; Siomi, H.; Adachi, A.; Oroszlan, S.; Maki, M.; Hatanaka, M. A region of basic amino-acid cluster in HIV-1 Tat protein is essential for trans-acting activity and nucleolar localization. Virus Genes 1989, 3, 99.
45.Ruben, S.; Perkins, A.; Purcell, R.; Joung, K.; Sia, R.; Burghoff, R.; Haseltine, W. A.; Rosen, C. A. Structural and functional characterization of human immunodeficiency virus tat protein. J. Virol. 1989, 63, 1.
46.Kao, S. Y.; Calman, A. F.; Luciw, P. A.; Peterlin, B. M. Anti-termination of transcription within the long terminal repeat of HIV-1 by tat gene product. Nature 1987, 330, 489.
47.Dayton, A. I.; Sodroski, J. G.; Rosen, C. A.; Goh, W. C.; Haseltine, W. A. The trans-activator gene of the human T cell lymphotropic virus type III is required for replication. Cell 1986, 44, 941.
48.Stevens, M.; De Clercq, E.; Balzarini, J. The regulation of HIV-1 transcription: molecular targets for chemotherapeutic intervention. Med. Res. Rev. 2006, 26, 595.
49.Ott, M.; Geyer, M.; Zhou, Q. The control of HIV transcription: keeping RNA polymerase II on track. Cell Host Microbe. 2011, 10, 426.
50.Wei, P.; Garber, M. E.; Fang, S. M.; Fischer, W. H.; Jones, K. A. A novel CDK9-associated C-type cyclin interacts directly with HIV-1 Tat and mediates its high-affinity, loop-specific binding to TAR RNA. Cell 1998, 92, 451.
51.Yamaguchi, Y.; Takagi, T.; Wada, T.; Yano, K.; Furuya, A.; Sugimoto, S.; Hasegawa, J.; Handa, H. NELF, a multisubunit complex containing RD, cooperates with DSIF to repress RNA polymerase II elongation. Cell 1999, 97, 41.
52.Wada, T.; Takagi, T.; Yamaguchi, Y.; Ferdous, A.; Imai, T.; Hirose, S.; Sugimoto, S.; Yano, K.; Hartzog, G. A.; Winston, F.; Buratowski, S.; Handa, H. DSIF, a novel transcription elongation factor that regulates RNA polymerase II processivity, is composed of human Spt4 and Spt5 homologs. Genes Dev. 1998, 12, 343.
53.Yamaguchi, Y.; Wada, T.; Watanabe, D.; Takagi, T.; Hasegawa, J.; Handa, H. Structure and function of the human transcription elongation factor DSIF. J. Biol. Chem. 1999, 274, 8085.
54.Price, D. H. P-TEFb, a cyclin-dependent kinase controlling elongation by RNA polymerase II. Mol. Cell. Biol. 2000, 20, 2629.
55.Kim, Y. K.; Bourgeois, C. F.; Isel, C.; Churcher, M. J.; Karn, J. Phosphorylation of the RNA polymerase II carboxyl-terminal domain by CDK9 is directly responsible for human immunodeficiency virus type 1 Tat-activated transcriptional elongation. Mol. Cell Biol. 2002, 22, 4622.
56.Gallego, J.; Varani, G. Targeting RNA with small-molecule drugs: therapeutic promise and chemical challenges. Acc. Chem. Res. 2001, 34, 836.
57.Athanassiou, Z.; Dias, R. L.; Moehle, K.; Dobson, N.; Varani, G.; Robinson, J. A. Structural mimicry of retroviral tat proteins by constrained β-hairpin peptidomimetics: ligands with high affinity and selectivity for viral TAR RNA regulatory elements. J. Am. Chem. Soc. 2004, 126, 6906.
58.Davidson, A.; Patora-Komisarska, K.; Robinson, J. A.; Varani, G. Essential structural requirements for specific recognition of HIV TAR RNA by peptide mimetics of Tat protein. Nucleic Acids Res. 2011, 39, 248.
59.Tamilarasu, N.; Huq, I.; Rana, T. M. Targeting RNA with peptidomimetic oligomers in human cells. Bioorg. Med. Chem. Lett. 2001, 11, 505.
60.Huq, I.; Wang, X.; Rana, T. M. Specific recognition of HIV-1 TAR RNA by a D-Tat peptide. Nat. Struct. Biol. 1997, 4, 881.
61.Simon, R. J.; Kania, R. S.; Zuckermann, R. N.; Huebner, V. D.; Jewell, D. A.; Banville, S.; Ng, S.; Wang, L.; Rosenberg, S.; Marlowe, C. K.; et al. Peptoids: a modular approach to drug discovery. Proc. Natl. Acad. Sci. U. S. A. 1992, 89, 9367.
62.Hamy, F.; Felder, E. R.; Heizmann, G.; Lazdins, J.; Aboul-ela, F.; Varani, G.; Karn, J.; Klimkait, T. An inhibitor of the Tat/TAR RNA interaction that effectively suppresses HIV-1 replication. Proc. Natl. Acad. Sci. U. S. A. 1997, 94, 3548.
63.Kesavan, V.; Tamilarasu, N.; Cao, H.; Rana, T. M. A new class of RNA-binding oligomers: peptoid amide and ester analogues. Bioconjug. Chem. 2002, 13, 1171.
64.Wang, X. H., I.; Rana, T. M. HIV-1 TAR RNA recognition by an unnatural biopolymer. J. Am. Chem. Soc. 1997, 119.
65.Tamilarasu, N. H., I.; Rana, T. M. . High affinity and specific binding of HIV-1 TAR RNA by a Tat-derived oligourea. J. Am. Chem. Soc. 1999, 121.
66.Bedford, M. T.; Clarke, S. G. Protein arginine methylation in mammals: who, what, and why. Mol. Cell. 2009, 33, 1.
67.Xie, B.; Invernizzi, C. F.; Richard, S.; Wainberg, M. A. Arginine methylation of the human immunodeficiency virus type 1 Tat protein by PRMT6 negatively affects Tat Interactions with both cyclin T1 and the Tat transactivation region. J. Virol. 2007, 81, 4226.
68.Frankel, A.; Yadav, N.; Lee, J.; Branscombe, T. L.; Clarke, S.; Bedford, M. T. The novel human protein arginine N-methyltransferase PRMT6 is a nuclear enzyme displaying unique substrate specificity. J. Biol. Chem. 2002, 277, 3537.
69.Calnan, B. J.; Tidor, B.; Biancalana, S.; Hudson, D.; Frankel, A. D. Arginine-mediated RNA recognition: the arginine fork. Science 1991, 252, 1167.
70.Fischer, R.; Fotin-Mleczek, M.; Hufnagel, H.; Brock, R. Break on through to the other side-biophysics and cell biology shed light on cell-penetrating peptides. Chem. Bio. Chem. 2005, 6, 2126.
71.Vives, E.; Brodin, P.; Lebleu, B. A truncated HIV-1 Tat protein basic domain rapidly translocates through the plasma membrane and accumulates in the cell nucleus. J. Biol. Chem. 1997, 272, 16010.
72.Conner, S. D.; Schmid, S. L. Regulated portals of entry into the cell. Nature 2003, 422, 37.
73.Khalil, I. A.; Kogure, K.; Akita, H.; Harashima, H. Uptake pathways and subsequent intracellular trafficking in nonviral gene delivery. Pharmacol. Rev. 2006, 58, 32.
74.Rothbard, J. B.; Jessop, T. C.; Lewis, R. S.; Murray, B. A.; Wender, P. A. Role of membrane potential and hydrogen bonding in the mechanism of translocation of guanidinium-rich peptides into cells. J. Am. Chem. Soc. 2004, 126, 9506.
75.Wender, P. A.; Galliher, W. C.; Goun, E. A.; Jones, L. R.; Pillow, T. H. The design of guanidinium-rich transporters and their internalization mechanisms. Adv. Drug. Deliv. Rev. 2008, 60, 452.
76.Calnan, B. J.; Biancalana, S.; Hudson, D.; Frankel, A. D. Analysis of arginine-rich peptides from the HIV Tat protein reveals unusual features of RNA-protein recognition. Genes Dev. 1991, 5, 201.
77.Delling, U.; Roy, S.; Sumner-Smith, M.; Barnett, R.; Reid, L.; Rosen, C. A.; Sonenberg, N. The number of positively charged amino acids in the basic domain of Tat is critical for trans-activation and complex formation with TAR RNA. Proc. Natl. Acad. Sci. U. S. A. 1991, 88, 6234.
78.Subramanian, T.; Govindarajan, R.; Chinnadurai, G. Heterologous basic domain substitutions in the HIV-1 Tat protein reveal an arginine-rich motif required for transactivation. EMBO J. 1991, 10, 2311.
79.Bahadur, R. P.; Zacharias, M.; Janin, J. Dissecting protein-RNA recognition sites. Nucleic Acids Res. 2008, 36, 2705.
80.Waksman, G.; Kominos, D.; Robertson, S. C.; Pant, N.; Baltimore, D.; Birge, R. B.; Cowburn, D.; Hanafusa, H.; Mayer, B. J.; Overduin, M.; Resh, M. D.; Rios, C. B.; Silverman, L.; Kuriyan, J. Crystal structure of the phosphotyrosine recognition domain SH2 of v-src complexed with tyrosine-phosphorylated peptides. Nature 1992, 358, 646.
81.Wender, P. A.; Mitchell, D. J.; Pattabiraman, K.; Pelkey, E. T.; Steinman, L.; Rothbard, J. B. The design, synthesis, and evaluation of molecules that enable or enhance cellular uptake: Peptoid molecular transporters. Proc. Natl. Acad. Sci. U. S. A. 2000, 97, 13003.
82.Paik, W. K.; Kim, S. Protein methylation: chemical, enzymological, and biological significance. Adv. Enzymol. Relat. Areas. Mol. Biol. 1975, 42, 227.
83.Blackwell, E.; Ceman, S. Arginine methylation of RNA-binding proteins regulates cell function and differentiation. Mol. Reprod. Dev. 2012, 79, 163.
84.Atherton, E.; Fox, H.; Harkiss, D.; Logan, C. J.; Sheppard, R. C.; Williams, B. J. Mild procedure for solid-phase peptide-synthesis- use of "fluorenylmethoxycarbonylamino-acids. J. Chem. Soc., Chem. Commun. 1978, 537.
85.Fields, G. B.; Noble, R. L. Solid phase peptide synthesis utilizing 9-fluorenylmethoxycarbonyl amino acids. Int. J. Pept. Protein Res. 1990, 35, 161.
86.Pace, C. N.; Vajdos, F.; Fee, L.; Grimsley, G.; Gray, T. How to measure and predict the molar absorption-coefficient of a protein. Protein Sci. 1995, 4, 2411.
87.Edelhoch, H. Spectroscopic determination of tryptophan and tyrosine in proteins. Biochemistry 1967, 6, 1948.
88.Garner, M. M.; Revzin, A. A gel electrophoresis method for quantifying the binding of proteins to specific DNA regions: application to components of the Escherichia coli lactose operon regulatory system. Nucleic Acids Res. 1981, 9, 3047.
89.Wang, J.; Huang, S. Y.; Choudhury, I.; Leibowitz, M. J.; Stein, S. Use of a polyethylene glycol-peptide conjugate in a competition gel shift assay for screening potential antagonists of HIV-1 Tat protein binding to TAR RNA. Anal. Biochem. 1995, 232, 238.
90.Puglisi, J. D.; Chen, L.; Blanchard, S.; Frankel, A. D. Solution structure of a bovine immunodeficiency virus Tat-TAR peptide-RNA complex. Science 1995, 270, 1200.
91.Marciniak, R. A.; Calnan, B. J.; Frankel, A. D.; Sharp, P. A. HIV-1 Tat protein trans-activates transcription in vitro. Cell 1990, 63, 791.
92.Lischwe, M. A.; Cook, R. G.; Ahn, Y. S.; Yeoman, L. C.; Busch, H. Clustering of glycine and NG,NG-dimethylarginine in nucleolar protein C23. Biochemistry 1985, 24, 6025.
93.Ziegler, A.; Seelig, J. Interaction of the protein transduction domain of HIV-1 Tat with heparan sulfate: binding mechanism and thermodynamic parameters. Biophys. J. 2004, 86, 254.
94.Nakase, I.; Takeuchi, T.; Tanaka, G.; Futaki, S. Methodological and cellular aspects that govern the internalization mechanisms of arginine-rich cell-penetrating peptides. Adv. Drug. Deliv. Rev. 2008, 60, 598.
95.Atherton, E.; Fox, H.; Harkiss, D.; Logan, C. J.; Sheppard, R. C.; Williams, B. J. A mild procedure for solid phase peptide synthesis: use of fluorenylmethoxycarbonylamino-acids. J. Chem. Soc., Chem. Commun. 1978, 537.
96.Richard, J. P.; Melikov, K.; Vives, E.; Ramos, C.; Verbeure, B.; Gait, M. J.; Chernomordik, L. V.; Lebleu, B. Cell-penetrating peptides. A reevaluation of the mechanism of cellular uptake. J. Biol. Chem. 2003, 278, 585.