Banca de DEFESA: Reynaldo Magalhães Melo

Uma banca de DEFESA de DOUTORADO foi cadastrada pelo programa.
STUDENT : Reynaldo Magalhães Melo
DATE: 30/10/2023
TIME: 09:00
LOCAL: Videoconferência
TITLE:

"Integration of top-down and bottom-up proteomics for the proteoform characterization of microorganism with medical and biotechnological relevance."

KEY WORDS:

Top-down proteomics, Bottom-up proteomics, Proteoforms, Amino acid production, Chagas disease, Corynebacterium glutamicum, Trypanosoma cruzi.

PAGES: 262
BIG AREA: Ciências Biológicas
AREA: Biologia Geral
SUMMARY:

Proteomics is a research field that employs methodologies capable of identifying, quantifying and characterizing proteins and their post-translational modifications (PTMs). There are two main proteomic approaches based on mass spectrometry (MS), known as bottom-up (BU) and top-down (TD). The former analyzes peptides derived from proteolytic digestion, and the latter directly examines intact proteins. Recently, the integration of these two approaches has enabled improvements in the identification and characterization of proteoforms, defined as different molecular forms produced by a single gene. This study applied BU and TD proteomic approaches to investigate Corynebacterium glutamicum, an important industrial workhorse for amino acid production, and Trypanosoma cruzi, the causative agent of Chagas disease. In C. glutamicum, the TD proteomic analysis revealed new proteoforms of OdhI, an important regulator of glutamate production, and mepB, a peptidase related to cell envelope metabolism. Additionally, quantitative BU proteomic analysis of C. glutamicum, comparing control and glutamate-induced production conditions, unveiled uncharacterized proteins involved in the early stages of this process. Further, a study was initiated to integrate BU and TD approaches for C. glutamicum under the same conditions, suggesting the influence of proteoforms related to nitrogen assimilation and branched-chain amino acid metabolism in the glutamate production process. Moreover, using the BU approach for PTM identification and subsequently employing this information to TD proteomics resulted in a significant increase in the number of identified proteoforms. In the case of T. cruzi, the BU approach was applied for quantitative proteomic analysis of axenic and intracellular amastigote forms, revealing a large number of regulated proteins between conditions. Among the regulated proteins were trans-sialidases, Leishmanolysin-like peptidases, and proteins associated with kDNA, suggesting the importance of these proteins in the replication process of T. cruzi. Lastly, the TD proteomic analysis was performed for T. cruzi epimastigotes, suggesting the identification of proteoforms caused by amino acid residue substitutions in proteins related with responses to environmental stress. The preliminary analysis also indicated methodologies that can be optimized for a better characterization of T. cruzi proteoforms, such as the identification of PTMs using the BU approach and the use of these data for proteoform identification via TD

COMMITTEE MEMBERS:
Externo à Instituição - AGENOR DE CASTRO MOREIRA DOS SANTOS JÚNIOR - FHB
Presidente - 404930 - CARLOS ANDRE ORNELAS RICART
Externa à Instituição - LYRIS MARTINS FRANCO DE GODOY - FIOCRUZ
Externo à Instituição - MAGNO RODRIGUES JUNQUEIRA - UFRJ
Interno - 2594909 - SEBASTIEN OLIVIER CHARNEAU