In-silico analysis of the novel Caenorhabditis elegans tropomyosin with poor actin affinity Público

Abstract

Tropomyosin is an actin-binding protein found in fungi and metazoans, which regulates actin filament stability and actomyosin contraction. In metazoans, multiple tropomyosin isoforms are expressed, and many of them are involved in diverse actin-dependent processes in an isoform-specific manner. In mammals, more than 40 tropomyosin isoforms are produced, and point mutations in several isoforms cause cardiovascular and skeletal muscle diseases. However, many tropomyosin isoforms remain poorly characterized. To investigate the biological significance of tropomyosin isoforms, we use the nematode Caenorhabditis elegans as a model organism. In C. elegans, the lev-11 gene is the sole tropomyosin gene that is essential for embryonic development, reproduction, and regulation of muscle contractility. The lev-11 gene is controlled by two separate promoters and extensive alternative splicing, and previous studies have demonstrated production of six tropomyosin isoforms. We identified a novel seventh exon, E7c, of the lev-11 gene and cloned a full-length cDNA encoding a novel low-molecular-weight tropomyosin isoform, LEV-11U that contains the E7c sequence. Interestingly, LEV-11U poorly bound to actin filaments in vitro, whereas other isoforms strongly bound to actin filaments. Here, I analyzed biophysical and biochemical properties of E7s of C. elegans tropomyosin isoforms. Fourier analysis showed that the 13^th-order acidic periodicity, which is an actin-adapted periodicity, of LEV-11U is weaker than that of LEV-11T. Sequence alignment showed that not only several periodic residues were not conserved in the E7c-encoded sequence but it also contained a unique substitution that violates the periodicity. Lastly, molecular dynamics simulations indicated that the E7c-encoded region poorly formed a stable coiled-coil structure. On the other hand, E7a and E7b formed unconventional interhelical bonds to stabilize their coiled-coil structures. These results indicate that E7c of the C. elegans lev-11 gene encodes a peptide sequence that is both biochemically and biophysically unique as compared with the equivalent sequences of other tropomyosin isoforms.

Table of Contents

Introduction 1

Methods 4

Results- Periodical analysis of the tropomyosin isoforms 7

Results- In silico structural analysis of the tropomyosin isoforms 9

Future directions 13

Figures 14

Appendix A: Steps to obtain coiled-coil centers 24

Appendix B: The full sequence of LEV-11U organized in the 13^th-order periodicity 26

References 27

About this Honors Thesis

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School	Emory College
Department	Mathematics
Degree	B.S.
Submission	Honors Thesis
Language	English
Research Field	Applied Mathematics
Palabra Clave	coiled-coil Caenorhabditis elegans (C. elegans) tropomyosin Fourier analysis alternative splicing molecular dynamics
Committee Chair / Thesis Advisor	Ono, Shoichiro, Emory University
Committee Members	Mayo, Talea, Emory University Shekhar, Shashank, Emory University

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