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Mistranslation and Quality Control of Aminoacyl-tRNA Synthetases

Han, Nien-Ching
http://rave.ohiolink.edu/etdc/view?acc_num=osu161842536059665

Abstract Details

2021, Doctor of Philosophy, Ohio State University, Microbiology.
The central dogma of biology is defined by the flow of genetic information from DNA to protein. In order to correctly interpret the information encoded in genomes, it is crucial for cells to maintain fidelity in DNA replication, transcription and translation. Among these three steps, translation was found to be the most erroneous, with the error frequency of one mistake every 1,000 to 10,000 codons translated. Errors in translation could occur at two different stages: aminoacylation and ribosomal decoding. In this thesis work, we focus on mistranslation which belongs in the first category. Aminoacylation is the reaction by which amino acids are attached onto tRNAs, forming aminoacyl-tRNAs. This process is catalyzed by aminoacyl-tRNA synthetases (aaRS) through two steps: amino acid activation and tRNA transfer. Due to the structural and chemical similarities shared by certain amino acids, it is sometimes difficult for the enzymes to distinguish non-cognate from cognate substrates, resulting in amino acid misactivation, which can potentially lead to mistranslation. Mistranslation is in general considered toxic because it may result in the production of aberrant or misfolded proteins, disrupting proteome homeostasis. To avoid mistranslation from occurring, some aaRSs have developed proofreading (editing) mechanisms, in which misactivated amino acids or mischarged tRNAs are hydrolyzed. Besides the endogenous editing activities of aaRS, additional trans-editing factors were identified in all domains of life, showing the importance of faithful aminoacylation. Non-proteinogenic amino acids (NPAs) are amino acids that are not naturally encoded in DNA. These substrates are often more problematic for translation integrity than their proteinogenic counterparts. One example is β-N-methylamino-L-alanine (BMAA), a neurotoxin generated by cyanobacteria that has been associated with diseases including amyotrophic lateral sclerosis (ALS) and Alzheimer’s disease (AD). Although BMAA has been hypothesized to enter the human proteome through mistranslation, questions remained on its mechanism of incorporation. We reported that BMAA is a substrate for human AlaRS (HsAlaRS), and is able to inhibit the enzyme's amino acid activation activity by acting as a competitive inhibitor. Additionally, we found that the endogenous editing function of HsAlaRS can be compromised by BMAA. Our results demonstrated that NPAs can disrupt the integrity of protein synthesis through different modes of action. Although editing is crucial in most cases, it was found that in many parasitic organisms the aaRS proofreading mechanisms are dispensable. For example, Mycoplasma species were reported with degenerated PheRS, ThrRS and LeuRS, which leads them to produce mischarged tRNAs, resulting in elevated amino acid misincorporation. It has been suggested that the adaptation of a promiscuous protein synthesis machinery may provide a gain of fitness for these organisms, but no evidence has been reported. In this thesis work, we characterized the amino acid activation domain of Mycoplasma mobile PheRS (MmPheRS) by steady-state activation kinetics using 6 different MmPheRS variants and three substrates: Phe, m-Tyr and Tyr. We revealed that although some mutations allowed the enzyme to discriminate cognate from non-cognate amino acids with higher specificities, it was at the cost of catalytic efficiency toward Phe. These results suggested that the ability to utilize available amino acids may be more important than the specificity for these host-dependent organisms, as they rely heavily on the nitrogen source the hosts provide.
Michael Ibba, Dr. (Advisor)
Kurt Fredrick, Dr. (Advisor)
Kou-San Ju, Dr. (Committee Member)
Juan Alfonzo, Dr. (Committee Member)
Biochemistry; Microbiology; Molecular Biology
Aminoacyl-tRNA synthetase; Translation quality control; Mistranslation; BMAA; AlaRS; PheRS

Recommended Citations

Citations

  • Han, N.-C. (2021). Mistranslation and Quality Control of Aminoacyl-tRNA Synthetases [Doctoral dissertation, Ohio State University]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=osu161842536059665

    APA Style (7th edition)

  • Han, Nien-Ching. Mistranslation and Quality Control of Aminoacyl-tRNA Synthetases. 2021. Ohio State University, Doctoral dissertation. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=osu161842536059665.

    MLA Style (8th edition)

  • Han, Nien-Ching. "Mistranslation and Quality Control of Aminoacyl-tRNA Synthetases." Doctoral dissertation, Ohio State University, 2021. http://rave.ohiolink.edu/etdc/view?acc_num=osu161842536059665

    Chicago Manual of Style (17th edition)

osu161842536059665
301
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