Avaliação dos produtos de expressão das regiões gênicas envolvidas na degradação do ácido úrico

Abstract

In hominids, the final product of purine catabolism is uric acid, unlike in other placental mammals where uric acid is degraded into (S)-allantoin through a series of reactions catalyzed by urate oxidase (uricase), HIU hydrolase (HIUase), and OHCU decarboxylase. In Homo sapiens, studies on the uric acid degradation pathway revealed that mutations in the uricase gene have led to reduced selective pressure for maintaining the gene regions of other enzymes in the pathway. The lack of expression of these enzymes can lead, in some situations, to the accumulation of uric acid. Despite the apparent absence of the proteins in H. sapiens, our analyses showed transcription of the gene regions for uricase (UOX), HIUase (URAHP), OHCU decarboxylase (URAD), and allantoicase (ALLC). While some transcripts of UOX and URAHP are considered non-productive, the genes URAD and ALLC are productive, meaning they produce protein-coding transcripts. In this study, literature data and public databases were analyzed to gather information on the products (RNA and protein) and expression profiles of the UOX, URAHP, URAD, and ALLC regions. This was followed by RNA-seq data analysis from case-control studies involving patients with conditions associated with hyperuricemia, to assess whether the expression of transcripts from these gene regions could contribute to the regulation of purine catabolism and the pathogenesis of diseases associated with uric acid accumulation. Since the initial hypothesis of the study was null and considering the potential presence of protein products from URAD and ALLC, evaluation was conducted on the possibility of producing active enzymes. Thus, alignments of the primary structure were performed, indicating that the protein product of URAD seemed more promising. Subsequently, modeling and evaluation of the tertiary structure were carried out, followed by expression in a heterologous system of recombinant H. sapiens OHCU decarboxylase. Upon obtaining the recombinant protein, enzymatic assays were performed, alongside characterization experiments using mass spectrometry, circular dichroism, and nuclear magnetic resonance, with the overall result indicating production of an active enzyme whose structure is consistent with other decarboxylases reported in the literature.