Characterization of DhHal3p: A moderately thermostable FMN-Binding Flavoprotein with biomedical potential from halotolerant yeast, Debaryomyces hansenii using partial structure prediction

Abstract

Enzymes within the CoA biosynthetic pathway are highlighted in the literature as promising targets for antimicrobial drugs, beyond their role in metabolism. In Saccharomyces cerevisiae, the model yeast, the Hal3 protein is a pivotal component of the PPCDC complex, a critical enzyme involved in the fourth step of Coenzyme A (CoA) biosynthetic pathway. Characterizing such proteins from extremophilic strains might present their substantial therapeutic potential. Therefore, the focus of the present study was to identify and characterize the putative DhHal3 gene and encoded protein from Debaryomyces hansenii, a halotolerant and teleomorph of commensal yeast, Candida famata. DhHal3 encoded a 559 amino acids peptide with a unique 48 amino acids aspartic acid-rich C-terminal domain. DhHal3p was identified as a flavoprotein (PFAM ID "PF02441") with conserved PLXANT and PXMNXXMW motifs, and H³⁴⁴ residue involved in FMN binding. Heterogeneously expressed 6xHistidine-
tagged DhHal3p appeared as ~73.37kDa protein on SDS-PAGE, exhibiting pyruvate decarboxylation activity (V₀ = 0.57 units/mL) in vitro. Thermo-profiling and circular dichroism (CD) analysis suggested DhHal3p is a moderately thermostable FMN-binding flavoprotein from D. hansenii. Docking simulations supported strong interactions between DhHal3p structure and FMN (binding energy = -4.04 kcal/mol). Further investigation into the functional characteristics of DhHal3p could yield pivotal insights into a variety of cellular processes, paving the way for therapeutic interventions.