Structural Features of Proteins Involved in Pfu RNase P or Recruitment of Viral Genomes to Human Chromatin
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2016
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ndltd-OhioLink-oai-etd.ohiolink.edu-osu14716217082021-12-20T05:37:19Z Structural Features of Proteins Involved in Pfu RNase P or Recruitment of Viral Genomes to Human Chromatin Crowe, Brandon L. Biochemistry Biophysics Structural biology NMR Protein protein interactions Transcription Factors Retroviral Integration Proteins perform important biological functions as part of large complexes with other macromolecules. However, biochemical studies have focused on individual proteins and domains due to the difficulty of studying large multi-subunit systems. This approach leaves a gap in our understanding between how the individual proteins function and their role in large macromolecular complexes. My research focused on bridging this gap by studying structural features proteins utilize to interact with other macromolecules, leading to their biological activity.I studied the interaction between RPP30 and Pop5, two protein subunits of Pyrococcus furiosus (Pfu) RNase P, an essential ribonucleoprotein complex required for 5'-maturation of tRNA. RPP30 and Pop5 are proteins, which interact and function as a pair in increasing the activity of the RNA component of RNase P. In chapter 2, I studied the RPP30-Pop5 complex in solution by gel filtration chromatography, dynamic light scattering and nuclear magnetic resonance (NMR). I found the RPP30-Pop5 complex is a heterotetrameric complex governed by hydrophobic packing, consistent with the crystal structure from a closely-related archaeon. In Appendix A, I used structural information from a variety of sources and molecular modeling to generate a tentative model of Pfu RNase P RNA. I also docked one copy of RPP21, RPP29, RPP30, and Pop5 with three copies of L7Ae to form a putative model of the holoenzyme. This structural model will be helpful for understanding and forming hypotheses on how the proteins modulate the structure and function of the RNA component. In Chapter 3, I investigated how the transcription factor Lens Epithelium Derived Growth Factor (LEDGF/p75) binds chromatin tightly enough to allow tethering of the pre-integration complex (PIC) of HIV-1. I first characterized the NMR solution structure of the PWWP domain from LEDGF/p75, and subsequently the regions on the PWWP domain used in binding to a peptide of histone protein H3 containing a tri-methylated lysine as well as double stranded deoxyribonucleic acid (DNA). I determined the Kd for both of these interactions and showed that while the interactions of each individual component with the PWWP domain is weak, the cumulative effect of binding both components at adjacent sites leads to tight binding of full nucleosomes.Lastly, for Chapter 4, I examined how the Bromodomain and Extraterminal domain (BET) family members bind the integrase of Murine Leukemia Virus (MLV) to tether it to host chromatin. I determined the structure of the complex between the extraterminal (ET) domain of Brd4 bound to a conserved 17-amino acid fragment of MLV integrase. I found that the binding involves the formation of an intermolecular three-stranded ß sheet between features that are unstructured on their own. The interaction is tight and governed by both hydrophobic packing and electrostatic interactions.Overall, my research has informed the structural features of proteins used in intermolecular complexes important for biological activity. 2016 English text The Ohio State University / OhioLINK http://rave.ohiolink.edu/etdc/view?acc_num=osu1471621708 http://rave.ohiolink.edu/etdc/view?acc_num=osu1471621708 unrestricted This thesis or dissertation is protected by copyright: all rights reserved. It may not be copied or redistributed beyond the terms of applicable copyright laws. |
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NDLTD |
| language |
English |
| sources |
NDLTD |
| topic |
Biochemistry Biophysics Structural biology NMR Protein protein interactions Transcription Factors Retroviral Integration |
| spellingShingle |
Biochemistry Biophysics Structural biology NMR Protein protein interactions Transcription Factors Retroviral Integration Crowe, Brandon L. Structural Features of Proteins Involved in Pfu RNase P or Recruitment of Viral Genomes to Human Chromatin |
| author |
Crowe, Brandon L. |
| author_facet |
Crowe, Brandon L. |
| author_sort |
Crowe, Brandon L. |
| title |
Structural Features of Proteins Involved in Pfu RNase P or Recruitment of Viral Genomes to Human Chromatin |
| title_short |
Structural Features of Proteins Involved in Pfu RNase P or Recruitment of Viral Genomes to Human Chromatin |
| title_full |
Structural Features of Proteins Involved in Pfu RNase P or Recruitment of Viral Genomes to Human Chromatin |
| title_fullStr |
Structural Features of Proteins Involved in Pfu RNase P or Recruitment of Viral Genomes to Human Chromatin |
| title_full_unstemmed |
Structural Features of Proteins Involved in Pfu RNase P or Recruitment of Viral Genomes to Human Chromatin |
| title_sort |
structural features of proteins involved in pfu rnase p or recruitment of viral genomes to human chromatin |
| publisher |
The Ohio State University / OhioLINK |
| publishDate |
2016 |
| url |
http://rave.ohiolink.edu/etdc/view?acc_num=osu1471621708 |
| work_keys_str_mv |
AT crowebrandonl structuralfeaturesofproteinsinvolvedinpfurnaseporrecruitmentofviralgenomestohumanchromatin |
| _version_ |
1723964994821816320 |