Use of TSAR, Thermal Shift Analysis in R, to Identify Metabolic Acid Molecules that Interact with HIV-1 Capsid Restricted; Files Only

Abstract

Thermal shift assay (TSA) is a simple, yet versatile technique used to screen for potential in vitro protein–ligand interactions. Here, we report a free, open-source software tool TSAR (Thermal Shift Analysis in R) to expedite and streamline the analysis of thermal shift data. The TSAR package incorporates three dynamic workflows to facilitate the analysis of TSA data derived either from individual experiments or large screens of chemical libraries. The pipelines aim to returns publication-ready graphics and processed summary statistics. TSAR offers dashboard-style graphic user interface (GUI) that enables easy use by non-programmers, simplifying TSA analysis while diversifying visualization. Accompanying the development and demonstration purposes of this software package, we screened two chemical libraries of vitamins and metabolic acids. Our method and data identify molecules interacting with the capsid protein (CA) of human immunodeficiency virus type 1 (HIV-1), where the hexameric CA interacts with numerous acids in vitro, many of which host potential biological significance given previous reports and subsequent studies. 

Table of Contents

1. INTRODUCTION ........................................................................................................................ 1

1.1 THEORETICAL BACKGROUND OF TSA ................................................................................... 2

2. LITERATURE REVIEW AND HYPOTHESIS .................................................................................... 4

2.1 COMPUTATIONAL SIGNIFICANCE ......................................................................................... 4

2.2 BIOLOGICAL HYPOTHESIS .................................................................................................... 5

3. METHODS ................................................................................................................................. 7

3.1 DISTRIBUTION AND PACKAGE INFORMATION ...................................................................... 7

3.2 PACKAGE STRUCTURE .......................................................................................................... 7

3.3 STATISTICS AND ANALYSIS .................................................................................................. 8

3.3.1 TSAR Algorithms ............................................................................................................ 8

3.3.2 Tm B Algorithm .............................................................................................................. 9

3.3.3 Tm D Algorithm ............................................................................................................. 11

3.3.4 TSAR Accuracy .............................................................................................................. 12

3.4 THERMAL SHIFT ASSAY (TSA).............................................................................................. 13

3.5 ENDOGENOUS REVERSE TRANSCRIPTION (ERT) ASSAY ....................................................... 15

3.6 CYTOTOXICITY TESTING .................................................................................................... 15

4. BIOLOGICAL DATA ................................................................................................................... 16

4.1 VITAMIN LIBRARY .............................................................................................................. 16

4.2 ORGANIC ACID METABOLITE LIBRARY ................................................................................ 18

5. BIOLOGY DATA DISCUSSION..................................................................................................... 20

5.1 UNDERSTANDING INTERACTIONS WITH FOLIC ACID ........................................................... 20

5.2 INHIBITORY EFFORTS OF GALLIC ACID ................................................................................ 21

6. SUPPLEMENT ........................................................................................................................... 23

6.1 ABBREVIATIONS .................................................................................................................. 23

6.2 SUPPLEMENTARY GRAPHS ................................................................................................... 25

7. REFERENCES ............................................................................................................................. 26 

About this Honors Thesis

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School	Emory College
Department	Quantitative Science
Degree	B.S.
Submission	Honors Thesis
Language	English
Research Field	Biology, Molecular Biology, Bioinformatics Health Sciences, Pharmacology
Keyword	Biology, Biostatistics
Committee Chair / Thesis Advisor	Stefan Sarafianos, Emory University Gordon Berman, Emory University
Committee Members	David Gorkin, Emory University

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