MUTUALISM IN NUCLEIC ACID/PEPTIDE DOMAIN ARRAYS: IMPLICATIONS FOR ORIGINS OF LIFE, NANOTECHNOLOGY, AND DISEASE Öffentlichkeit

Abstract

The intricate connections between nucleic acids and proteins in the cell produces a network of spatially and temporally regulated mutualisms. These mutualisms likely preceded cellular life, and are essential to the central dogma. The ribosome, a conglomerate of proteins and ribosomal RNAs is conserved and regarded as the Darwinian threshold for cellular life. Less complex mutualisms are expected to have preceded the ribosome and supported the first cellular networks in protocells. The stabilization and propagation of nucleic acids with homogeneous 5'-3' linkages, a necessary prerequisite to the organization of a replicative system, was likely mediated by proteins or small peptides that served to protect nucleic acids from the harsh prebiotic environment. Peptide assemblies, which may have formed prebiotically upon concentration of peptides as short as two amino acids in a discrete area, are being explored as scaffolds for specific nucleic acid elongation. The use of peptide assemblies as scaffolds is also exploited in nanotechnology where the production of peptide hydrogels for tissue engineering continues. The complementarity of nucleic acids is manipulated in the construction of DNA origami, where its digital-like interactions are fine-tuned for the development of responsive systems. The diversity of DNA secondary structures and their context dependence extends these efforts. Guanine quadruplexes, which demonstrate efficient electron transfer are being explored as wires in bionanocircuitry. Combination of the scaffolding properties of peptide assemblies and the diverse complementary folding landscape of DNA, highlights the fabrication of artificial mutualisms. Extant mutualisms between DNA and RNA binding proteins and their targets in the cell, are responsible for the spatiotemporal regulation of cellular information flow. RNA is processed in membraneless organelles known as ribonucleoprotein granules (RNP). The liquid-liquid phase transitions that characterize the assembly and disassembly of RNP granules is mediated by RNA binding proteins with low complexity domains (LCD). Disruption of transient interactions between LCDs, or seeding of infectious domains by reversible LCD amyloids, is considered a contributor to altered ribostasis in protein misfolding diseases where deposition of RNA at disease lesions has not yet been explained. Here we explore these diverse mutualisms through structural characterization of a novel RNA/peptide co-assembly.

Table of Contents

Table of Contents

Chapter 1: Looked at Life from Both Sides Now..............................................................................1

From up and down and still somehow.......................................................................................1

Biopolymer diversity..................................................................................................................3

Functional assemblies...............................................................................................................7

From both sides now.................................................................................................................9

Conclusions: toward molecular mutualisms............................................................................12

Chapter 2: Altering nanotube groove morphology to promote nucleic acid elongation..................20

INTRODUCTION.....................................................................................................................20

RESULTS................................................................................................................................22

Self-assembly of reactive-neutral nanotubes..........................................................................23

Self-assembly of groove-modified peptide nanostructures.....................................................27

Synthesis of modified adenine nucleotides.............................................................................38

Nucleic acid elongation on nanotubes.....................................................................................40

CONCLUSIONS......................................................................................................................49

METHODS...............................................................................................................................51

Chapter 3: DNA/peptide chimeras: manipulation of mutualisms for functional applications..........58

INTRODUCTION.....................................................................................................................58

RESULTS................................................................................................................................62

Synthesis of DNA/Peptide conjugates.....................................................................................62

Spectroscopic identification of guanine quadruplexes............................................................66

GQPC/Ac-KLVIIAG-NH2 co-assembly....................................................................................69

Guanine quadruplexes form during conical GQPC/peptide co-assembly...............................74

Specific guanine quadruplex recognition..........................................................................76

Creating responsive hydrogels................................................................................................79

Assembly of nucleic acid/peptide conjugates (NAPC)............................................................81

CONCLUSIONS......................................................................................................................86

METHODS...............................................................................................................................87

Chapter 4: Design and global architecture of nucleic acid/peptide co-assemblies........................96

INTRODUCTION.....................................................................................................................96

RESULTS................................................................................................................................98

Sampling of peptide congeners for nucleic acid co-assembly.................................................98

Assembly of homogeneous nucleic acid/peptide nanostructures..........................................103

Particle formation in RNA/peptide nanostructure assembly..................................................107

Cooperative binding of nucleic acids to peptide assemblies.................................................109

Temperature affects global co-assembly architecture...........................................................112

Microscopy confirms co-assembly of peptides and nucleic acid...........................................115

Evaluating the electrostatic contribution to RNA/peptide co-assembly.................................117

RNA/peptide co-assemblies remain dynamic........................................................................121

Co-assembly of dsDNA and pep-KG/RG..............................................................................123

CONCLUSIONS....................................................................................................................125

METHODS.............................................................................................................................126

Chapter 5: Passivation of the cross-b interface by nucleic acids.................................................134

INTRODUCTION...................................................................................................................134

RESULTS..............................................................................................................................134

RNA/peptide co-assemblies maintain cross-b architecture...................................................135

Co-assemblies form homogeneous anti-parallel, in-register b-sheet monolayers................137

DQF-DRAWS identifies parallel component of peptide assembly.........................................145

Nucleic acids passivate the cross-b monolayer interfaces....................................................146

Synthesis and characterization of a ¹³C/³¹P calibration standard..........................................149

Molecular dynamics simulations of DNA/pep-KG co-assemblies..........................................154

Co-assembly structural model...............................................................................................158

CONCLUSIONS....................................................................................................................159

METHODS.............................................................................................................................160

Chapter 6: A b /RNA co-assemblies and disease etiology............................................................170

INTRODUCTION...................................................................................................................170

RESULTS..............................................................................................................................171

A b 40 and 42 fibril surfaces are ideal for templating nucleic acids.........................................172

Co-assembly of A b 40 and 42 with RNA................................................................................173

Visualization of co-assembly by fluorescence microscopy....................................................178

Circular Dichroism identifies order in co-assemblies.............................................................180

Powder x-ray diffraction of co-assemblies.............................................................................182

CONCLUSIONS....................................................................................................................184

METHODS.............................................................................................................................186

Chapter 7: Concluding remarks...................................................................................................192

Development of chimeras for bionanotechnology.................................................................193

Engineering homogeneous nucleic acid/peptide co-assemblies...........................................194

Comprehensive models for RNA processing and disease etiology.......................................195

About this Dissertation

Rights statement

• Permission granted by the author to include this thesis or dissertation in this repository. All rights reserved by the author. Please contact the author for information regarding the reproduction and use of this thesis or dissertation.

School	Laney Graduate School
Department	Biological and Biomedical Sciences
Subfield / Discipline	Biochemistry, Cell & Developmental Biology
Degree	PhD
Submission	Dissertation
Language	English
Research Field	Chemistry, Biochemistry Nanotechnology Chemistry, General
Stichwort	Ribonucleoprotein granule Protein Misfolding Co-assembly
Committee Chair / Thesis Advisor	Lynn, David, Emory University
Committee Members	Koval, Michael H, Emory University Deal, Roger, Emory University Matsumura, Ichiro, Emory University Ortlund, Eric, Emory University

Zuletzt geändert

Primary PDF

Thumbnail	Title	Date Uploaded	Actions
	MUTUALISM IN NUCLEIC ACID/PEPTIDE DOMAIN ARRAYS: IMPLICATIONS FOR ORIGINS OF LIFE, NANOTECHNOLOGY, AND DISEASE ()	2018-08-28 16:24:49 -0400	Press to Select an action Download

Supplemental Files

Thumbnail	Title	Date Uploaded	Actions